J. Biomedical Science and Engineering, 2010, 3, 65-77
doi:10.4236/jbise.2010.31010 Published Online January 2010 (http://www.SciRP.org/journal/jbise/
Published Online January 2010 in SciRes. http://www.scirp.org/journal/jbise
Phosphatidylinositol transfer proteins: sequence motifs
in structural and evolutionary analyses
Gerald J. Wyckoff1, Ada Solidar2, Marilyn D. Yoder3
1Division of Molecular Biology and Biochemistry, University of Missouri-Kansas City, Kansas City, USA;
2Vassa Informatics, Kansas City, USA;
3Division of Cell Biology and Biophysics, University of Missouri-Kansas City, Kansas City, USA.
Email: wyckoffg@umkc.edu
Received 30 June 2009; revised 28 September 2009; accepted 30 September 2009.
Phosphatidylinositol transfer proteins (PITP) are a
family of monomeric proteins that bind and transfer
phosphatidylinositol and phosphatidylcholine be-
tween membrane compartments. They are required
for production of inositol and diacylglycerol second
messengers, and are found in most metazoan organ-
isms. While PITPs are known to carry out crucial
cell-signaling roles in many organisms, the structure,
function and evolution of the majority of family
members remains unexplored; primarily because the
ubiquity and diversity of the family thwarts tradi-
tional methods of global alignment. To surmount this
obstacle, we instead took a novel approach, using
MEME and a parsimony-based analysis to create a
cladogram of conserved sequence motifs in 56 PITP
family proteins from 26 species. In keeping with pre-
vious functional annotations, three clades were sup-
ported within our evolutionary analysis; two classes
of soluble proteins and a class of membrane-associat-
ed proteins. By, focusing on conserved regions, the
analysis allowed for in depth queries regarding pos-
sible functional roles of PITP proteins in both intra-
and extra- cellular signaling.
Keywords: Protein Evolution; Structural Domain;
Phylogenetics; Sequence Motif
Phosphatidylinositol transfer proteins (PITP) are mono-
meric, lipid-binding proteins that bind and transfer
phosphatidylinositol (PtdIns) and phosphatidylcholine
(PtdCho) between membrane compartments (see reviews
by: [1,2,3,4,5,6]. Inositol lipids have specialized func-
tions in the regulation of eukaryotic cells, providing a
source of second messengers and acting as signaling
molecules. Monomeric phospholipids have extremely
low solubilities and negligible spontaneous transfer rates
between membranes, necessitating protein factors to
shield them in the aqueous environment of the cell. Al-
most all phospholipid exchange activity within the eu-
karyotic cytosol is accomplished by three groups of pro-
teins: PITP, phosphatidylcholine transfer protein (PCTP),
or sterol carrier protein 2. PITP-domain proteins, which
are the focus of this study, are found in five classes of
proteins: three are soluble and cytosolic, and two are
membrane-associated proteins [1,2] (Figure 1).
Soluble PITPs are found in virtually all metazoan or-
ganisms. These are approximately 77% identical at the
amino-acid level. Two isoforms, PITP and PITP, exist
in mammals; they are highly conserved with about 98%
sequence identity at the amino acid level. The proteins
bind one molecule of PtdIns or PtdCho and are typically
about 32 kDa in mass.
PITP-like domains are also detected in the retinal de-
generation B (rdgB) class of proteins (see reviews [6,7]).
These 160-170 kDa proteins are membrane-associated
proteins first identified in Drosophila melanogaster, as
mutations in these genes lead to retinal degeneration.
The rdgBs contain an
Figure 1. Gene structure of PITP-domain proteins. Grey cyl-
inder is PITP-like domain, white cylinder is the FFAT region of
ER binding, the grey box is the DDHD region, and the white
box is the LSN2 region. The the human pitpnm1 protein, the
FFAT region is residues 360-365, the DDHD region is 686-879,
and the LSN2 domain is residues 1022-1152 [6].
66 G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
SciRes Copyright © 2010 JBiSE
(a) (b) (c)
Figure 2. Cartoon representation of rat PITP complexed to PtdCho. (a). Structural domains of the class I PITPs displayed on a
representative PITP (PDB:1T27). The lipid binding core is shown in blue, the regulatory loop in green, and the C-terminal region
in red. The PtdCho molecule is in transparent spheres with standard CPK coloring; (b) Conserved sequence motifs mapped onto
the same structure. The four conserved sequence motifs; m4, m9, m13, m14, and m23, representing branch points in the clado-
gram shown in Figure 3 are in purple and are annotated. The remaining sequence motifs are in gold, regions not present in con-
served sequence motifs are colored grey; (c). Image B is rotated 90º counterclockwise around a vertical axis.
N-terminal PITP-like domain (42% sequence identity to
PITP isoforms), a FFAT sequence motif [2,8], a DDHD
metal binding domain (which has calcium-binding capa-
bilities in the human proteins [9]), and a LNS2 domain.
The FFAT motif is a short sequence containing two
phenylalanines in an acidic tract that targets the protein
to the endoplasmic reticulum [8]. The DDHD domain of
180 residues may be involved in metal binding, but the
function of this domain is unknown [6]. The LNS2 do-
main is believed to be involved in protein: protein inter-
actions, and in the human homologues is a protein tyro-
sine kinase 2 (PYK2) binding domain [9]. Two homo-
logues of rdgB are found in most mammalian genomes
studied to date and are usually called 1) rdgB(I), rdgBI,
or PITPnm1 and 2) rdgB(II), rdgBII, or PITPnm2. In
Drosophila, rdgB is believed to function in the termina-
tion of phototransduction and in the establishment and
maintenance of rhodopsin levels in photoreceptor cells
[10,11]. In humans, PITPnm1 has widespread tissue dis-
tribution and can rescue fly rdgB mutant phenotypes [1];
whereas PITPnm2 has a neuronal-specific expression
pattern and is unable to functionally rescue fly rdgB
mutants [12]. Although Drosophila rdgB possesses the
capability to transfer PtdCho and PtdIns in vitro [10,11],
the PITP-domains of rdgB and PITP are not function-
ally interchangeable [10].
An additional class of PITP-like proteins, rdgB, has
been identified in mice, humans, and Drosophila. These
are 38 kDa, soluble proteins that have sequence similar-
ity most comparable to the N-terminal region of rdgB-
class proteins. It shares approximately 40% sequence
identity with PITP/. The purified protein has been
shown to possess in vitro PtdIns transfer capabilities
Plants and fungi generally do not contain a sequence
homologue to PITP, but do possess a functional analog
referred to as Sec14p in yeast systems. Sec14p is ap-
proximately the same size as PITP, and although there is
no detectable amino acid similarity between the two
proteins, temperature-sensitive mutants of yeast Sec14p
are rescued by rat PITP and PITP [15,16]. Likewise,
Sec14p can successfully substitute for PITP in the PITP-
dependent reconstitutions studied to date [17,18,19,20].
Interestingly, the slime mold Dicytostelium discoideum
has been shown to contain not only homologues to PITP,
called DdPITP1 and DdPITP2, but also a Sec14p rela-
tive, called DdSec14 [21].
The experimentally determined three-dimensional
structures of rat PITP-PtdCho [22] and PITP-PtdCho
[23], human PITP-PtdIns [24], and the apo form of
mouse PITP [25] have been reported. The PITP struc-
tures share little resemblance to the crystal structure of
yeast Sec14p [26]. The PITP structure is composed of
three regions (Figure 2(a)): a lipid-binding core, a regu-
latory loop, and the C-terminal region. The lipid-binding
core of PITP-PtdCho shares a fold with the steroido-
genic acute regulatory protein-related transfer (START)
domain [22] first observed in human MLN64 [27]. It has
been proposed that the START domain may be a common
fold adapted for binding lipid molecules. PCTP is a
START-domain protein based on structure and sequence
G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
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Figure 3. Phylogenetic tree for 56 PITP family proteins. A strict con-
sensus tree was derived from the presence/absence of sequence motifs
as well as the motif sequence, analyzed via parsimony as described in
the text. The blue lines show which sequence motifs are uniformly
gained within each clade on the tree.
identity, whereas PITP is not considered to be a START-
domain protein due to a lack of sequence identity.
Mutations and gene-knockout studies provide insight
into the functions of PITP. In mice, a mutation in the
PITP gene causes the vibrator phenotype, which is
characterized by a progressive-action tremor, degenera-
tion of brain stem and spinal cord neurons, and juvenile
death. PITPβ does not compensate for loss of PITP
[40]. Furthermore, in mice, embryonic stem cells defi-
cient in PITP or PITP reveal differences in physio-
logical function between the two isoforms. PITP defi-
ciency leads to catastrophic failure early in embryonic
development, and the protein is therefore posited to have
an essential housekeeping role in the cell [29]. In con-
trast, PITP-deficient embryonic stem cells are not
compromised in growth or in bulk phospholipid metabo-
lism; however, PITP is required for neonatal survival.
PITP deficiency affects regulation of phospholipid
transport in the ER, endocrine pancreas function, and
glycogen metabolism, due to compromised lipid absorp-
tion, homeostatic problems, and severe hypoglycemia
Protein similarity is usually detected by conservation
of function as revealed by biochemical analysis, sequence
68 G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
SciRes Copyright © 2010 JBiSE
Figure 4. Sequence motifs mapped to the structure of rat PITP:PtdCho.
The sequence motifs present in class I PITPs are shown in a rainbow spec-
trum of colors from blue to red and from the N- to the C-terminus. Se-
quence motifs are annotated ‘m1’ to m25’, the PtdCho molecule is in
transparent spheres with standard CPK coloring. The lipid binding core
contains sequence motifs 1-2, 4-7, 9-10, 13-15, and 17-25. The regulatory
loop contains sequence motifs 15, 17, and 18-20. The C-terminal region
contains part of sequence motif 24 and all of motif 25.
similarity as detected by amino acid pattern rec ognition,
or by structural similarity as detected by X-ray crystal-
lography or NMR spectroscopy. The PITP/rdgB proteins
possess intriguing patterns of similarity. PITP has func-
tional similarity to Sec14p, but lacks sequence or struc-
ture similarity. PITPs have sequence similarity to rdgB
and both possess the hallmark ability to transfer phos-
pholipids between the protein and membranes. PITP has
been shown to have structural similarity to the START
domains, even though it does not share amino acid se-
quence similarity. Here, we have used a comprehensive
evolutionary analysis to synthesize information from
sequence analysis and structural comparisons. This ap-
proach leads to a cohesive understanding of evolutionary,
structural, and functional relationships of the PITP/rdgB
protein families, and may suggest that a re-analysis of
PITP protein naming conventions is overdue.
2.1. Identification of Sequences
Rat PITP (gi:8393962) was used as a query sequence
in a BLASTp (version 2.2.11, database versions as of
May 8, 2005) [31] search. The 134 returned sequences
were manually culled to remove duplicated or signifi-
cantly fragmented sequences, reducing the number to 60
sequences in 26 species (Table 5). The final data set
contained both cytosolic PITP and rdgBs of approxi-
mately 270-330 residues each, as well as the membrane-
associated rdgB/PITPnm proteins, of approximately
1250 residues. Several proteins were determined to be
splice variants of existing proteins in the tree; they are
annotated in all figures and tables.
2.2. Phylogenetic Analysis
Unaligned proteins were processed using the MEME
program [32] (implemented in the Wisconsin Package,
version 10.4) [33], using the “zero or more” setting,
which permits anywhere from zero instances of a block
to extreme repetition of blocks, thereby permitting the
generation of the most robust matrix of sequence motifs
possible. The phylogenetic parsimony tree for the se-
quence motifs was made by PAUP* (version 4.0b) [34].
The consensus tree was imported into MacClade (v 4.08)
[35] for further character-state analysis. Character-state
changes were traced along tree clades and unambiguous
changes were examined. A database of sequence motifs
and proteins was created to facilitate this study and the
G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
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Table 1. Conserved sequence motif matrix of PITP and rdgB proteins, PITP domain only.
tables from it are available upon request.
In this phylogenetic analysis, the tree root was desig-
nated by the branch that incorporated the most sequence
motif additions and that led to the most consistent char-
acter-state changes along the tree. That is, the basal prot-
eins that rooted the tree had the fewest sequence motifs
character-state changes within the tree with this root
present. This rooting was borne out by the consistency and
was utilized. This rooting suggested the fewest character
state changes and least amount of homoplasy within the
tree. The tree root consisted of small PITP proteins from
Plasmodium, Giardia, and Encephalitozoon. Sequence
motif 14 defined this branch point, proteins from these
species lacked the motif, and all other proteins possessed
Sequence motif identifier
ing # 1 2 3 4 5 6 7 8 9 1011121314151617181920 21 22 232425
1 1 1 0 0 1 1 1 0 0 10000001010 1 0 0 10
2 1 1 0 0 1 1 1 0 0 10000001110 1 1 0 10
3 1 1 0 0 1 1 1 0 0 10000001110 0 0 0 10
4 1 1 0 0 1 1 0 0 0 10000001110 0 0 0 00
59 0 1 0 0 1 1 0 0 0 10000000000 0 0 0 10
60 0 1 0 0 0 1 0 0 0 00000000000 0 0 0 00
56 1 1 0 1 1 1 0 0 1 10001101010 1 0 0 10
15-16 1 1 1 0 1 1 0 1 0 10001010110 1 0 0 10
17-19, 25,
28-29 1 1 1 0 1 1 1 1 0 10101010110 1 0 0 10
20 0 0 1 0 1 1 1 1 0 10101010110 1 0 0 10
21-24 1 1 1 0 1 1 1 1 0 10101010110 1 0 0 11
26 0 0 0 0 1 1 1 1 0 10101010110 1 0 0 10
27 1 1 1 0 1 1 1 1 0 10101001110 1 0 0 10
30 1 1 1 0 1 1 1 1 0 10101001110 1 0 0 00
57 1 1 0 0 1 1 1 1 0 10001100110 1 0 0 10
5-7 1 1 1 0 1 1 1 1 0 11001001110 1 0 0 10
8 0 1 1 0 1 1 1 1 0 11001001110 1 0 0 10
9 0 1 1 0 1 1 1 1 0 11001001010 1 0 0 10
10 1 1 1 0 1 1 1 1 0 11001001110 1 0 0 00
11 1 1 1 0 1 1 1 1 0 11001001010 1 0 0 10
12 1 1 1 0 1 1 1 1 0 11000001110 1 0 0 10
58 1 1 0 1 1 1 0 0 0 10001001010 1 0 0 10
34 1 1 0 1 1 1 1 0 1 10001101110 1 1 1 10
53 1 1 0 1 1 1 1 0 1 10011101110 0 0 1 11
54 0 1 0 1 1 1 0 0 1 10011101110 0 0 1 11
42-51 1 1 0 1 1 1 1 0 1 10011101111 1 1 1 11
40-41, 52 0 1 0 1 1 1 1 0 1 10011101111 1 1 1 11
55 1 1 1 0 1 1 1 0 0 10001001110 1 0 0 11
32 0 1 0 1 1 1 0 0 1 10001001010 1 0 0 10
33 1 1 0 1 1 1 0 0 1 10001001010 1 0 0 10
70 G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
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Table 2. Conserved sequence motif matrix of rdgB proteins, C-terminal of the PITP-domain.
Table 3. Gene and protein nomenclature for proteins utilized in this study.
the conserved sequence motif.
3.1. Characterization of PITP-Like Proteins
An initial approach to characterizing the PITP family
was relatively standard. Rat PITP was used as the seed
for a BLAST search targeting relatively distant protein
relatives. BLAST results were imported to Pileup [33] to
generate a “first pass” global alignment. Under a variety
of different conditions, including weighting end gaps,
not weighting end gaps, utilizing low gap opening and
extension penalties, high-road and low-road alignment
options, alignments were very weak, with notable cases
where locally homologous segments identified by BLAST
were not aligned in the global matrix. The derived nature
of many of the proteins in the PITP family was already
apparent; sequence motifs appeared in some classes but
not others, rendering standard alignment and analysis
methods ineffective. A novel approach was required to
identify and analyze the large, divergent PITP family.
3.2. Identification of Conserved Sequence Motifs
When a high-confidence global alignment in Pileup
proved impossible, unaligned proteins were processed
using MEME [32] which identifies conserved sequence
motifs in proteins. The program makes identifications
based on prior probabilities of amino acid occurrence.
Because it performs ungapped determinations, no preced-
ing global alignment of sequence motifs was necessary,
families with highly derived members. Of the 46 cons-
erved sequence motifs detected by MEME, 25 fell
Sequence motif identifier
king # 26 27 28 29 30 31 32 333435363738394041 42 43 44 45 46
15 0 0 0 0 1 1 0 0 11 1 1 1 1 10 1 0 0 0 0
16 0 0 0 0 1 1 0 0 11 1 1 1 1 10 1 0 0 0 1
26, 28 1 1 1 1 1 1 1 1 11 1 1 1 1 11 1 1 1 0 0
18 1 1 1 1 1 1 1 0 11 1 1 1 1 11 1 1 0 0 0
20 0 1 1 0 1 1 0 0 11 1 1 1 1 11 1 1 0 0 0
19 1 0 1 1 1 1 0 0 11 1 1 1 1 11 1 0 0 0 0
23 1 1 1 1 1 1 1 1 11 1 1 1 1 11 1 1 1 1 0
24 1 1 0 1 1 1 1 1 11 1 1 1 1 11 1 1 1 1 0
25 1 1 1 1 0 1 0 0 11 1 1 1 1 11 1 1 0 0 0
27 1 1 1 1 1 1 1 1 11 1 1 1 1 11 1 1 0 0 0
29 1 1 1 0 0 0 0 0 11 1 1 1 1 11 0 0 0 0 0
30 1 0 1 1 1 1 1 0 11 1 1 0 0 00 0 0 0 0 0
PITP PITP  rdgBβ rdgB(1) rdgB(2)
Gene name
Pitpna (mouse)
Pitpn (rat)
Pitpna (human)
Pitpnb (mouse)
Pitpnb (rat)
Pitpnb (human)
pitpnc1 (human)
pitpnm1 (mouse)
rdgB1 (fly)
pitpnm2 (mouse)
NIR3 rdgB2
Gene sym-
Pitpna (mouse)
Pitpn (rat)
Pitpna (human)
Pitpnb (mouse)
Pitpnb (rat)
Pitpnb (human)
pitpnc1 (human)pitpnm1 (mouse) pitpnm2 (mouse)
Gene ali-
Pitpn, vib1A
Vib1B Rd9; RdgB; DRES9;
mpt-1; Pitpnm
NIR3; rdgB2;
rdgB 2;
rdgBβ ; rdgB  
1; PITP, cyto-
plasmic 1*
PITP mem-
brane-associated 1
PITP mem-
brane-associated 2
gene sym-
G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
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Table 4. Conserved sequence motifs in the PITP-domain.
Motif Identi-
secondary structure element,
Sequence rangeb:
Motif sequenceb:
1 sheet 1 (3-11) 2-9 2-VLLKEYRV-9
1 1-8 1-MLIKEYHI-8
1 1-8 1-MLLKEYRI-8
2 helix A (14-33) 12-24 PVSVDEYQVGQLY
3 - --------
3 24-31 MIQKKSRE
3 24-31 MISKHSHE
4 helix A (14-33) 26-36 VAEASKNETGG
4 - -----------
4 - -----------
5 sheet 2 (39-49) 37-48 GEGVEVLVNEPY
6 sheet 3 (55-64) 55-64 GQYTHKIYHL
7 helix B (70-75) 66-73 SKVPTFVR
7 204-211 AKIEQFIH
7 198-205 TRVEQFVH
8 - -----------
9 - -----------
9 - -----------
10 sheet 4 (84-91) 87-96 AWNAYPYCRT
11 - --------
11 - --------
11 97-104 YTCSFLPK
12 - --------
12 98-105 RYTCPFVE
12 - --------
13 sheet 5 (94-100) 99-106 TNEYMKED
13 - --------
13 - --------
14 sheet 6 (108-117) 107-120 FLIKIETWHKPDLG
15 helix C (132-137) 123-134 ENVHKLEPEAWK
15 - ------------
15 - ------------
16 - -----------------------
16 - -----------------------
17 sheet 7 (138-142) 137-144 EAVYIDIA
17 - --------
17 134-141 EVCFIDIA
18 153-160 DYKAEEDP
18 152-159 EYKAEEDP
18 149-156 YYKESEDP
19 163-173 FKSIKTGRGPL
19 162-172 YHSVKTGRGPL
19 159-169 FKSEKTGRGQL
20 helix E (177-183) 175-183 PNWKQELVN
20 - ---------
20 - ---------
21 sheet 8 (191-201) 190-206 MCAYKLVTVKFKWWGLQ
72 G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
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22 helix F (206-232) 207-214 NKVENFIH
22 - --------
22 - --------
23 helix F (206-232) 217-224 ERRLFTNF
23 - --------
23 - --------
24 helix F (206-232) 225-244 HRQLFCWLDKWVDLTMDDIR
25 helix G (240-261) 245-253 RMEEETKRQL
25 243-252 ALEEETARML
25 - ----------
26 - --------
26 294-301 PPGPDASP
26 269-276 RSAPSSAP
aSecondary structure nomenclature for rat PITP as defined in [22], parenthesis indicate the complete range of the secondary structure
bSequences and residue numbering refer to human proteins, PITP is gi:5453908, tracking #38; PITPnm1 is gi:18490106, tracking #21;
PITPNC1 is gi:32307140, tracking #7. PITPNC1 is also called rdgB.
within PITP-like domains.
3.3. Phylogenetic Analysis
With sequence motifs identified, two character-state ma-
trices were created. One contained character-state infor-
mation on the presence or absence of detected motifs,
and one contained aligned amino acid data (Tables 1 and
2). These matrices were utilized for the creation of a
character state phylogenetic tree via maximum parsi-
mony. The overall shape of the tree was determined by
the presence/absence matrix, with the amino acid align-
ment data utilized to determine internal clade structures.
Gaps, i.e., missing sequence motifs were considered a
new state, and thus, the difference between having a
specific set of amino acids and not having an amino acid
could served as an informative character state.
Twelve most parsimonious trees of differing structures
were identified by PAUP, but the strict consensus tree
structure suggests the likely evolutionary history of this
intriguing family of proteins (Figure 3). The tree re-
vealed that membrane-associated proteins are derivatives
of a group of PITP-like genes, and that many genes pre-
viously thought to be related to PITP and PITP via se-
quence homology fell outside of clades that contain ex-
clusively PITP or PITP genes.
3.4. Identification of Clades Within the
Phylogenetic Tree
In order to be cladistically sound, nomenclature must be
hierarchical; all proteins in the tree were “PITP-like.”
The tree lends itself to consideration of the PITP-like
family consisting of three large divisions (the classifica-
tion nomenclature proposed by Allen-Baume [1] is
adapted here): Class I comprised the soluble PITPs,
Class IIA were the membrane-associated rdgB proteins
(which contain additional domains C-terminal to the
PITP-like domain), and Class IIB were the soluble
rdgB proteins. Further subdivisions along cladistic
lines allowed for the naming of the Alpha clade and the
Beta clade within Class I, and an ancestral group of
PITP-like proteins, predominantly from protista, that
occured in several clades rooting the tree. Table 3 lists
the nomenclature commonly utilized for the genes and
proteins in this study, as well as the HUGO-approved
3.5. Examination of Conserved Sequence Motifs
To understand the functional shifts that have occurred,
examination of some of the sequence motifs in each
proposed class was necessary. The amino acid sequence
of each motif in the human proteins PITP (Class I),
PITPNM1 (Class IIA), and PITPNC1, also called rdgB
(Class IIB) are given in Table 4.
Class I: These proteins are cytosolic and contain se-
quence motifs 1-2, 4-7, 9-10, 13-15, and 17-25 (Figure
4). This class was distinguished from the Class II divi-
sions by sequence motif 8, which was unique to the
Class IIA and IIB clades, and sequence motif 4, which
was unique to the non-protista Class I PITPs. A striking
observation about sequence motif 8 was that it appears
to be an amino acid sequence overlap with sequence
motif 9, which defines subnodes within Class I. In the
structure of rat PITP, the residues defined by sequence
motif 9 incorporate helix B, which is a putative mem-
brane insertion helix [25]. There was no unique con-
served sequence motif that distinguishes the functionally
divergent alpha and the beta clades.
Class IIA: These proteins are membrane-associated
and typically are 160 kDa. In this analysis they were
distinguished by the unique presence of sequence motifs
34-40, which consisted of amino acids residues 831-844,
856-866, 899-920, 937-951, 993-1015, 1022-1038, and
1040-1086 in the human PITPnm1 protein. The division
had two major subnodes, one consisting of non-mamma-
G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
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Table 5. Proteins used in phylogenetic analysis.
aindicated as a predicted protein, bindicated as a hypothetical or a putative protein
lian proteins and containing the canonical rdgB protein
from D. melanogaster, and the other subnode containing
the mammalian sequences PITPNM1 and PITPNM2.
There were 21 conserved sequence motifs outside the
gi # Tracking # Species common Protein designations
55243798 19 Anopheles gambiae mosquito
48135931 20 Apis mellifera honeybee
39590557 16 Caenorhabditis briggsaaenematode
39590636 33 Caenorhabditis briggsaaenematode CB09751 b (NCBI-COG: PITP)
17556182 32 Caenorhabditis elegans
17554244 15 Caenorhabditis elegans nematode PITP DDHD (NCBI-COG: PITP)
48059855 58 Caenorhabditis elegans nematode Y71G12B.17 b (NCBI-COG: PITP)
57091327 40 Canus familiaris dog similar to PITP a
41055500 49 Danio rerio zebrafish PITP 
41055576 51 Danio rerio zebrafish similar to PITP
28422482 52 Danio rerio zebrafish
8307957 56 Dictyostelium discoideumslime moldPITP 2
8307955 57 Dictyostelium discoideumslime moldPITP 1
24641869 17 Drosophila melanogasterfruit fly CG11111-PB, isoform B, rdgB
62484257 12 Drosophila melanogasterfruit fly CG17818-PA, rdgBβ
7300495 35 Drosophila melanogasterfruit fly CG5269-PA PITP, vib
20151901 34 Drosophila melanogasterfruit fly SD01527p, vib
54642914 18 D.pseudoobscura fly GA10766-PA, Dpse\GA10766
19170839 60 Encephalitozoon cuniculi PITP
50758480 41 Gallus gallus chicken similar to PITP a
50756343 31 Gallus gallus chicken similar to PITP, membrane-associated 2 a
50755397 13 Gallus gallus chicken similar to rdgBβ a
53134209 47 Gallus gallus chicken
50757849 8 Gallus gallus chicken similar splicing variant rdgBβ a
29250063 59 Giardia lamblia similiar to D. discoideum PITP1
18490106 21 Homo sapiens human PITPNM1
24308237 27 Homo sapiens human PITPNM2
5453908 38 Homo sapiens human PITP
6912594 43 Homo sapiens human PITPβ
32307140 7 Homo sapiens human PITP, rdgBβ 1
6679337 36 Mus musculus mouse PITP
9790159 45 Mus musculus mouse PITPβ 
22003862 5 Mus musculus mouse rdgB 
6679339 23 Mus musculus mouse PITPNM1
47124324 25 Mus musculus mouse PITPNM2
2137007 39 Oryctolagus cuniculus rabbit PITP
55660937 42 Pan troglodytes chimp PITP  a
55644759 54 Pan troglodytes chimp similar to PITP a
55639157 30 Pan troglodytes chimp similar to PITP membrane-associated 2 a
55636463 24 Pan troglodytes chimp similar to PITP membrane-associated 1 a
56493706 2 Plasmodium berghei PITP b
56509884 4 Plasmodium chabaudi protein b
23619433 3 Plasmodium falciparum PITP b
23485539 1 Plasmodium yoelii yoelii PITP 2
55731914 46 Pongo pygmaeus orangutan protein b
56605814 22 Rattus norvegicus rat PITPnm, PI membrane-associated a
62658898 26 Rattus norvegicus rat similar KIAA1457, PITPnm2a
8393962 37 Rattus norvegicus rat PITP, PITPn
16758568 44 Rattus norvegicus rat PITP 
62657241 6 Rattus norvegicus rat rdgB 
56756428 55 Schistosoma japonicum Unknown, similiar to vib (fly)
47228496 53 Tetraodon nigroviridis pufferfish
47228470 28 Tetraodon nigroviridis pufferfish
47223953 9 Tetraodon nigroviridis pufferfish
47206861 14 Tetraodon nigroviridis pufferfish
47226436 29 Tetraodon nigroviridis pufferfish
47937798 48 Xenopus laevis frog MGC84500 proteinb
49256286 10 Xenopus laevis frog MGC84224 protein
62860160 11 Xenopus tropicalis frog protein LOC549393 b
38512077 50 Xenopus tropicalis frog PITPβ
74 G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
SciRes Copyright © 2010 JBiSE
PITP domain, with sequence motifs 34-37 observed in
all analyzed proteins in this class. In human pitpnm1, the
FFAT domain was found at the end of sequence motif 27,
with a derived motif sequence of EFFDCLD. The
DDHD region contained sequence motifs 30-35, which
comprises 35% of the defined DDHD range, and the
LSN2 region contained sequence motifs 39 and the
N-terminal portion of sequence motif 40, which is 23%
of this region. Lev, et al. defined six, short hydrophobic
regions based on hydrophathy plots that were originally
postulated to be transmembrane domains [9] then later to
mediate membrane association [7]. None of the six re-
gions fell in a conserved sequence motif identified in
this study On the other hand, sequence motif 26 ap-
peared in multiple repeats throughout the non-PITP do-
main in sequence motifs 29, 32-33, and 44-45. This is a
Ser rich motif of eight amino acids, the significance of
this observation is currently unclear.
Class IIB: The proteins in this class are historically
referred to as rdgBs and are monomeric, cytosolic pro-
teins of ~330 amino acids. They were distinguished by
the unique presence of sequence motif 11, which con-
sists of residues 97-104 in the human PITPNC1 se-
quence. Sequence motif 11 is interesting; although it was
only present in proteins in this division, it overlaped in
sequence with motif 12 and motif 13. Sequence motif 12
was only observed in class IIA, whereas sequence motif
13 was unique to Class I, alpha and beta clades. Another
notable feature of this division was PITP1 from D. dis-
coideum. It grouped outside the other rdgBproteins  
and rooted the class IIA clade.
4.1. Phylogenetics
Ocaka et al [36] provide an extensive analysis of the
chromosomal location of the PITPN genes in humans
with a phylogenetic tree rooted with C. elegans PITP as
the chosen outgroup. The analysis presented here differs
in two respects. First, the original gene-duplication event
leading to the evolution of soluble and mem-
brane-associated PITP proteins likely occurred not early
in animal evolution, but instead was initiated in some
protists. D. discoideum has been shown to have two
genes, pitA and pitB, and expression has been demon-
strated for both proteins, PITP1 and PITP2 [21]. The
phylogeny shown here indicates that PITP1 is a class IIB
protein, perhaps representative of an ancestral precursor
to class IIA proteins. Second, data presented here indi-
cate that the Class IIB proteins are derived from Class I
proteins, and the membrane-associated Class IIA pro-
teins are derived from the class IIB proteins. Previous
cladograms and dendrograms have indicated different
derivization patterns. Ocaka et al [36] indicate that
mammalian PITPNC1 (Class IIB in the present analysis)
shares a most recent common ancestor (MRCA) with
PITPNA and PITPNB; more so than with any of the
PITPNMs. Fullwood et al. present a dendrogram in
which the membrane-associated PITPs and the soluble
PITPs split from a shared derivation from Class IIB pro-
teins (rdgB). An increased sample size, an unbiased
tree root, and the use of conserved sequence motifs for
global alignment in the present study lead to a different
interpretation of the evolution of this protein family.
The phylogenetic tree produced here is indicative of
the evolutionary history of these sequence motifs, and
not of the proteins as a whole. This is important, because
in some proteins, large sequences with no known se-
quence motif patterns are observed (such as in the Plas-
modium yoelli “PITP1” protein) that may be the result of
gene conversion, unequal crossing-over, or repeat ex-
pansion during the evolution of specific proteins. Such
large insertions and deletions in protein sequence are
difficult to utilize in any phylogenetic analysis; they are
purely derived characters that are phylogenetically un-
informative. However, the existence of these derived
sequences suggests that either the function or regulation
of proteins in this family has changed dramatically, in-
dicative of the type of specific adaptation often seen
within protein families.
4.2. Functional Shifts in PITP Proteins
Alignment and subsequent phylogenetic analysis of di-
vergent members of protein families is a problem com-
pounded by shifts in primary, secondary, and tertiary
structure that occur during protein evolution and func-
tional diversification. Global alignments of complete
protein coding regions often fail when functional units
(such as regulatory regions, transmembrane domains,
DNA-binding domains, protein-protein interaction do-
mains, etc.) are gained or lost over evolutionary time.
This issue complicates the otherwise straightforward
phylogenetic analysis of related proteins. Additional
complications arise in organismal surveys in which en-
tire genomes are sequenced and annotated by automated
algorithms based on homology. BLAST and FASTA
searches find protein regions with high local identity,
which can suggest functional similarity. However, out-
side of these identified regions, major changes can occur
in protein function that obscure identity and make global
protein alignment difficult or impossible. Without cor-
responding functional data, these genes are often named
inconsistently. For example, a gene found in one species
with locally high homology to a protein region in an-
other species is often named “Similar to…” or “Species
X homologue of Species Y protein”. Because BLAST is
a local homology search tool, it is inadequate for identi-
fication and annotation of species orthologs. For this
reason, many of the genes identified in species genome
projects are insufficiently annotated; phylogenetic
analysis of all proteins labeled as “Phosphatidylinositol
transfer proteins, Beta” for example will yield a spurious
G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
SciRes Copyright © 2010 JBiSE
alignment and therefore a misleading tree. As demon-
strated in this study, analysis of the PITP/rdgB family
was complicated by both issues. It should be noted that
full-length sequences produce similar phylogenetic trees
for alignments of many related sets of proteins (all Class
I or Class II proteins, for example).
The resultant phylogenetic tree reveals that the gen-
eral naming convention for the PITP family needs to be
critically examined. Many of the proteins in this family
cannot be named exclusively by local sequence identity,
as their functions appear to be significantly altered from
previously studied proteins. Many annotated genes are
named in a way that might not be truly indicative of their
function. In other words, sequence homology in local
segments has obscured the greater functional diversifica-
tion within this family. For example, although the mem-
brane-associated proteins (Class IIB) are shown to be
derived and monocladistic, the tree root appears to be
within the PITP family, suggesting that many other func-
tions are derived characteristics from this single-domain
protein. At best, this tree demonstrates the need for bet-
ter functional and structural classification of PITP-like
proteins before final assignment of their names.
An additional feature of the strict consensus tree is the
hypothesis it suggests about the evolution of PITP pro-
tein family members. First, no PITP proteins are found
within prokaryotes or within yeast. Yeast have an analo-
gous enzyme, Sec14p, but it appears to have an inde-
pendent evolutionary origin. The slime-mold and Plas-
modium sequences represent the most primitive species
in this PITP family tree. In contrast, the multiplication of
forms within mammals is impressive. Humans, mice,
and rats have well-characterized genomes and each have
a variety of PITP family-member proteins.
4.3. Conserved Sequence Motifs
The use of conserved sequence motifs in a cladistic
analysis of the PITP family highlights several regions for
consideration. Two regions are of particular interest: one
containing sequence motifs 7, 8, and/or 9, and the other
containing sequence motifs 11, 12, or 13. These two
regions form loops at the surface of the protein near the
ends of the acyl chains of the bound phospholipid (Fig-
ure 2(b) and 2(c)). Sequence motif 7 is the only motif
present that is not found in the same primary sequence
space in the three main protein classes. In human PITP,
the amino acid sequence is at residues 66-73, while it
occurs at residues 204-211 and 198-205 in human
PITPNM1 and PITPNC1, respectively. Sequence motif 8
is not present in class I PITPs, and partially overlaps
with sequence motif 9, which is exclusive to the al-
pha/beta clades of class I. Sequence motif 7 in the class I
PITP's partially overlaps with sequence motif 8 of class
IIA and IIB. Sequence motif 7 contains the putative helix
insertion loop proposed to anchor PITP to the mem-
brane during lipid exchange [25]. It is tempting to specu-
late that this helix has adapted a different conformation
or perhaps has migrated across the lipid-binding cavity
to near residues 200-208 relative to the PITP struc-
The second region of interest contains sequence mo-
tifs 11, 12, or 13. These three motifs are present in ap-
proximately the same space in primary sequence, but
each is unique to the three family classes. Sequence mo-
tif 11 is unique to class IIB, whereas Sequence motif 12
is unique to class IIA, and sequence motif 13 is unique
to class I. The role of this loop in the structure or the
function of any class of PITP is currently unexplored.
A concern with the novel phylogenetic analysis tech-
nique used in this study is the proportion of the sequence
data actually used in the evolutionary analysis and
whether amino acids that have been structurally and
functionally shown to affect phospholipid transfer and
signal transduction capabilities are included in the
alignment matrices. In human PITP, 214 of the 270
amino acids (79.3%) are included in the conserved se-
quence motifs. Most site-directed mutagenic analysis of
the role of specific residues in the phospholipid transfer,
specificity, and PLC reconstitution capabilities have
been done on human or rat PITP and PITP. Residues
in rat PITP that have been studied and shown to play a
functional role in PITPs include Thr59, Lys61, Glu86,
Asn90, Tyr103, and the double Trp pair at 203-204
[37,38]. These residues are all in conserved sequence
motifs. Thr59 and Lys 61 map to sequence motif 6,
which is the only sequence motif present in all se-
quences in this study. Interestingly, sequence motif 9,
containing Glu 86, is present only in Class I proteins.
Tyr103 has been shown to diminish PLC reconstitution
without affecting phospholipid transfer. This residue is a
Tyr only in the Class I proteins, and is a conservative
substitution to a Phe in the class IIA and class IIB pro-
teins. Examination of these amino acids and sequence
motifs from an evolutionary perspective enables a more
thorough understanding of their importance in shaping
PITP function and structure.
4.4. Functional Specialization across Evolution
Functional shifts within the PITP family are difficult to
assess in light of the relative paucity of experimental
analysis of different family members. In rat, PITPs are
broadly expressed and have been detected in at lease 20
tissues [39]. Unigene expression profiles (http://www.
ncbi.nlm.nih.gov/entrez/query.fcgi?db=unigene) for ma-
mmalian species indicate a slightly more general and
ubiquitous expression for the alpha isoform than the beta
isoform. Mouse PITP -/- embryonic stem cells are em-
bryonically lethal, indicating essential functions in cell
viability [29]. In contrast, mice deficient in PITP de-
velop normally to term, but fail to thrive neonatally [30].
76 G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
SciRes Copyright © 2010 JBiSE
These observations and the EST expression profiles
support the argument that PITP isoforms are not redun-
dant, and have different functions [29]. One implication
of the consensus tree is that, given that the ancestral
proteins have putative roles in cellular function, e.g.,
PITP2 in Dictyostelium, they may be more widely ex-
pressed than the more derived family members, which
appear to have roles only in specific tissues or at specific
developmental times. PITPnm1 in mice, for example, is
highly expressed primarily in the retina, with weaker
expression in the brain and some suggestion (from rat
data) of central nervous system expression. Other studies
in humans showed PITPnm2 and PITPnm3 expression in
brain and retina.
The phylogenetic analysis described here has demon-
strated the utility of the use of conserved sequence mo-
tifs in detecting evolutionary relationships among pro-
teins in large and/or diverse functions. This method uses
short conserved sequence motifs instead of single char-
acter amino acids. The presence and absence of motifs
becomes an important data point. The use of sequence
motifs in phylogenetic analysis as described here should
be applicable to the analysis of a wide range of protein
families, particularly large, diverse families where re-
shuffling of domains occurred over evolutionary time.
The authors wish to acknowledge Lee Likins, Christine Malcom, Justin
Paschall, and Ming Yang for comments and assistance. Funding for
this study was provided in part by a University of Missouri Research
Board grant and NIH R15HD055668-01A1 (to GJW).
[1] Allen-Baume, V., Segui, B. and Cockcroft, S. (2002)
Current thoughts on the phosphatidylinositol transfer
protein family. FEBS Letters, 531, 74-80.
[2] Cockcroft, S. and Carvou, N. (2007) Biochemical and
biological functions of class I phosphatidylinositol trans-
fer proteins. Biochim. Biophys. Acta, 1771, 677-691.
[3] Hsuan, J. and Cockcroft, S. (2001) The PITP family of
phosphatidylinositol transfer proteins. Genome Biol., 2,
[4] Routt, S.M. and Bankaitis, V.A. (2004) Biological func-
tions of phosphatidylinositol transfer proteins. Biochem.
Cell Biology, 82, 254-262.
[5] Thomas, G.M.H. and Pinxteren, J.A. (2000) Phosphati-
dylinositol transfer proteins: One big happy family or
strangers with the same name? Mol. Cell Biol. Res.
Comm., 4, 1-9.
[6] Wirtz, K.W.A. (2006) Phospholipid transfer proteins in
perspective. FEBS Lett., 580, 5436-5441.
[7] Trivedi, D. and Padinjat, R. (2007) RdgB proteins: func-
tions in lipid homeostasis and signal transduction. Bio-
chim. Biophys. Acta, 1771, 692-699.
[8] Loewen, C.J., Roy, A. and Levine, T.P. (2003) A con-
served ER targeting motif in three families of lipid bind-
ing proteins and in Opi1p binds VAP. EMBO J. 22,
[9] Lev, S., Hernandez, J., Martinez, R., Chen, A., Plowman,
G. and Schlessinger, J. (1999) Identification of a novel
family of targets of PYK2 related to Drosophila retinal
degeneration B (rdgB) protein. Mole. Cell. Biol., 19,
[10] Milligan, S.C., Alb, J.G. Jr., Elagina, R. B., Bankaitis, V.
A. and Hyde, D.R. (1997) The phosphatidylinositol
transfer protein domain of Drosophila retinal degenera-
tion B protein is essential for photoreceptor cell survival
and recovery from light stimulation. J. Cell Biol., 139,
[11] Vihtelic, T.S., Goebl, M., Milligan, S., O'Tousa, J.E. and
Hyde, D.R. (1993) Localization of Drosophila retinal
degeneration B, a membrane associated phosphatidy-
linositol transfer protein. J. Cell Biol., 122, 1013-1022.
[12] Lu, C., Vihtelic, T.S., Hyde, D.R. and Li, T. (1999) A
neuronal-specific mammalian homolog of the Drosophila
retinal degeneration B gene with expression restricted to
the retina and dentate gyrus. J. Neurosci., 19, 7317-7325.
[13] Fullwood, Y., dos Santos, M. and Hsuan, J.J. (1999)
Cloning and characterization of a novel human phos-
phatidylinositol transfer protein, rdgBb. J. Biol. Chem.,
274, 31553-31558.
[14] Takano, N., Owada, Y., Suzuki, R., Sakagami, H., Shi-
mosegawa, T. and Kondo, H. (2003) Cloning and char-
acterization of a novel variant (mM-rdgB b1) of mouse
M-rdgBs, mammalian homologs of Drosophila retinal
degeneration B gene proteins, and its mRNA localization
in mouse brain in comparison with other M-rdgBs. J
Neurochem, 84, 829-839.
[15] Skinner, H.B., Alb ,J.G.Jr., Whitters, E.A., Helmkamp, G.
M.Jr. and Bankaiitis V. A. (1993) Phospholipid transfer
activity is relevant to but not sufficient for the essential
function of the yeast SEC14 gene product. EMBO, 12,
[16] Tanaka, S. and Hosaka, K. (1994) Cloning of a cDNA
encoding a second phosphatidylinositol transfer protein
of rat brain by complementation of the yeast sec14 muta-
tion. J. Biochem (Tokyo), 115, 981-984.
[17] Cunningham, E., Tan, S.K., Swigart, P., Hsuan, J.,
Bankaitis, V. and Cockcroft, S. (1996) The yeast and
mammalian isoforms of phosphatidylinositol transfer
protein can all restore phospholipase C-mediated inositol
lipid signaling in cytosol-depleted RBL-2H3 and HL-60
cells. Proc. Natl. Acad. Sci. USA, 93, 6589-6593.
[18] Hay, J.C. and Martin, T.F.J. (1993) Phosphatidylinositol
transfer protein is required for ATP-dependent priming of
Ca2+-activated secretion. Nature, 366, 572-575
[19] Jones, S.M., Alb, J.G.J., Phillips, S. E., Bankaitis and
V.A., Howell, K.E. (1998) A phosphatidylinositol
3-kinase and phosphatidylinositol transfer protein act
synergistically in formation of constitutive transport
vesicles from the trans-Golgi network. J. Biol. Chem.
273, 10349-10354.
[20] Ohashi, M., de Vries, K.J., Frank, R., Snoek, G.,
Bankaitis, V., Wirtz, K. and Huttner, W.B. (1995) A role
for phosphatidylinositol transfer protein in secretory
vesicle formation. Nature, 377, 544-547.
[21] Swigart, P., Insall, R., Wilkins, A. and Cockcroft, S.
(2000) Purification and cloning of phosphatidylinositol
transfer proteins from Dictyostelium discoideum: homo-
G. J. Wyckoff et al. / J. Biomedical Science and Engineering 3 (2010) 65-77
SciRes Copyright © 2010
logues of both mammalian PITPs and Saccharomyces
cerevisiae Sec14p are found in the same cell. Biochem. J.,
347, 837-843.
[22] Yoder, M.D., Thomas, L.M., Tremblay, J.M., Oliver, R.
L., Yarbrough, L.R. and Helmkamp, G.M.Jr. (2001)
Structure of a multifunctional protein: mammalian phos-
phatidylinositol transfer protein complexed with phos-
phatidylcholine. J. Biol. Chem., 276, 9246-9252.
[23] Vordtriede, P.B, Doan, C.N., Tremblay, J.M., Helmkamp,
G.M.J. and Yoder, M.D. (2005) Structure of PITPb in
complex with phosphatidylcholine: Comparison of struc-
ture and lipid transfer to other PITP isoforms. Biochem-
istry, 44, 14760-14771.
[24] van Tiel, C.M., Schouten, A., Snoek, G.T., Gros, P., Wirtz,
K.W.A. (2002) The structure of phosphatidylinositol
transfer protein a reveals sites for phospholipid binding
and membrane association with major implications for its
function. FEBS, 531, 69-73.
[25] Schouten, A., Agianian, B., Westerman, J., Kroon, J.,
Wirtz, K.W.A. and Gros, P. (2002) Structure of apo-
phosphatidylinositol transfer protein a provides insight
into membrane association. EMBO J., 21, 2117-2121.
[26] Sha, B., Phillips, S.E., Bankaitis, V.A. and Luo, M. (1998)
Crystal structure of the Saccharomyces cerevisiae phos-
phatidylinositol transfer protein. Nature, 391, 506-510.
[27] Romanowski, M.J., Soccio, R.E., Breslow, J.L. and Bur-
ley, S.K. (2002) Crystal structure of the Mus musculus
cholesterol-regulated START protein 4 (StarD4) contain-
ing a StAR-related lipid transfer domain. Proc. Natl.
Acad. Sci. USA, 99, 6949-6954.
[28] Hamilton, B.A., Smith, D.J., Mueller, K.L., Kerrebrock,
A.W., Bronson, R.T., van Berkel, V., Daly, M.J.,
Kruglyak, L, Reeve, M.P., Nemhauser, J.L., Hawkins, T.
L. Rubin, E.M. and Lander, E.S. (1997) The vibrator
mutation causes neurodegeneration via reduced expres-
sion of PITPa: positional complementation cloning and
extragenic suppression. Neuron, 18, 711-722.
[29] Alb, J.G.Jr., Phillips, S.E., Rostand, K., Cui, X., Pinx-
teren, J., Cotlin, L., Manning, T, Guo, S, York, J.D.,
Sontheimer, H., Collawn, J.F., Bankaitis, V.A. (2002)
Genetic ablation of phosphatidylinositol transfer protein
function in murine embryonic stem cells. Mol. Biol. Cell,
13, 739-754.
[30] Alb, J.G.J., Cortese, J.D, Phillips, S.E., Albin, R.L., Nagy,
T.R., Hamilton, B.A. and Bankaitis, V.A. (2003) Mice
lacking phosphatidylinositol transfer protein-a exhibit
spinocerebellar degeneration, intestinal and hepatic stea-
tosis, and hypoglycemia. J. Biol. Chem. 278, 33501-
[31] Altschul, S.F., Gish, W., Miller, W., Myers, E.W., Lipman,
D.J. (1990) Basic local alignment search tool. J. Mol.
Biol. 215, 403-410.
[32] Bailey, T.L. and Elkan, C.P. (1994) Fitting a mixture
model by expectation maximization to discover motifs in
biopolymers. Proceedings of the Second International
Conference on Intelligent Systems for Molecular Biology.
AAAI Press, Menlo Park, CA, 28-36.
[33] Accelrys, I. (2005) Wisconsin Package v.10.3. Accelrys,
Inc, San Diego, CA.
[34] Swofford, D.L. (2003) PAUP*. Phylogenetic analysis
using parsimony and other methods. Version 4. Sinauer
Associates, Sunderland, Massachusetts.
[35] Maddison, D.R. and Maddison, W.P. (2005) MacClade
v.4.08. Sinauer Associates, Sunderland, Massachusetts.
[36] Ocaka, L., Spalluto, C., Wilson, D.I, Hunt, D.M., Halford,
S. (2005) Chromosomal localization, genomic organiza-
tion and evolution of the genes encoding human phos-
phatidylinositol transfer protein membrane- associated
(PITPNM) 1, 2 and 3. Cytogenet Genome Res., 108,
[37] Alb,J.G. Jr., Gedvilaite, A., Cartee, R.T., Skinner, H.B.,
Bankaitis, V.A. (1995) Mutant rat phosphatidylinosi-
tol/phosphatidylcholine transfer proteins specifically de-
fective in phosphatidylinositol transfer: Implications for
the regulation of phospholipid transfer activity. Proc.
Natl. Acad. Sci. USA, 92, 8826-8830.
[38] Tilley, S.J., Skippen, A., Murray-Rust, J., Swigart, P.M.,
Stewart, A., Morgan, C.P., Cockcroft, S. and McDonald,
N.Q. (2004) Structure-function analysis of phosphatidy-
linositol transfer protein alpha bound to human phos-
phatidylinositol. Structure, 12, 317-326.
[39] Venuti, S.E. and Helmkamp, G.M.J. (1988) Tissue distri-
bution, purification and characterization of rat phos-
phatidylinositol transfer protein. Biochim Biophys Acta,
946, 119-128.
[40] Lev, S. (2004) The role of the Nir/rdgB protein family in
membrane trafficking and cytoskeleton remodeling. Exp.
Cell Res., 297, 1-10.