Truncations of the C-Terminus Have Different Effects on the Conformation and Activity of Phosphatidylinositol Transfer Protein†

Biochemistry ◽  
1996 ◽  
Vol 35 (38) ◽  
pp. 12526-12531 ◽  
Author(s):  
Paul A. Voziyan ◽  
Jacqueline M. Tremblay ◽  
Lynwood R. Yarbrough ◽  
George M. Helmkamp
Keyword(s):  
Transfer Protein ◽  
C Terminus ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

scholarly journals Differential expression of a C-terminal splice variant of phosphatidylinositol transfer protein β lacking the constitutive-phosphorylated Ser262 that localizes to the Golgi compartment

2006 ◽  
Vol 398 (3) ◽  
pp. 411-421 ◽  
Author(s):  
Clive P. Morgan ◽  
Victoria Allen-Baume ◽  
Marko Radulovic ◽  
Michelle Li ◽  
Alison Skippen ◽  
...  
Keyword(s):  
Amino Acids ◽  
Rat Liver ◽  
Splice Variant ◽  
Single Gene ◽  
Hl60 Cells ◽  
Transfer Protein ◽  
Golgi Localization ◽  
C Terminus ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

Mammalian PITPβ (phosphatidylinositol transfer protein β) is a 272-amino-acid polypeptide capable of transferring PtdIns, PtdCho and SM (sphingomyelin) between membrane bilayers. It has been reported that Ser262 present in the C-terminus of PITPβ is constitutively phosphorylated and determines Golgi localization. We provide evidence for the expression of an sp (splice) variant of PITPβ (PITPβ-sp2) where the C-terminal 15 amino acids of PITPβ-sp1 are replaced by an alternative C-terminus of 16 amino acids. PITPβ-sp1 is the product of the first 11 exons, whereas PITPβ-sp2 is a product of the first 10 exons followed by the twelfth exon – exon 11 being ‘skipped’. Both splice variants are capable of PtdIns and PtdCho transfer, with PITPβ-sp2 being unable to transport SM. PITPβ is ubiquitously expressed, with the highest amounts of PITPβ found in HL60 cells and in rat liver; HL60 cells express only PITPβ-sp1, whereas rat liver expresses both sp variants in similar amounts. In both cell types, PITPβ-sp1 is constitutively phosphorylated and both the PtdIns and PtdCho forms of PITPβ-sp1 are present. In contrast, PITPβ-sp2 lacks the constitutively phosphorylated Ser262 (replaced with glutamine). Nonetheless, both PITPβ variants localize to the Golgi and, moreover, dephosphorylation of Ser262 of PITPβ-sp1 does not affect its Golgi localization. The presence of PITPβ sp variants adds an extra level of proteome complexity and, in rat liver, the single gene for PITPβ gives rise to seven distinct protein species that can be resolved on the basis of their charge differences.

scholarly journals Deletion of 24 amino acids from the C-terminus of phosphatidylinositol transfer protein causes loss of phospholipase C-mediated inositol lipid signalling

1997 ◽  
Vol 324 (1) ◽  
pp. 19-23 ◽  
Author(s):  
Simon PROSSER ◽  
Robert SARRA ◽  
Philip SWIGART ◽  
Andrew BALL ◽  
Shamshad COCKCROFT
Keyword(s):  
Amino Acids ◽  
Phospholipase C ◽  
Size Exclusion ◽  
Lipid Transfer ◽  
Transfer Protein ◽  
C Terminus ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

Phosphatidylinositol transfer protein α (PITPα) is a 32 kDa protein of 270 amino acids that is essential for phospholipase C-mediated phosphatidylinositol bisphosphate hydrolysis. In addition, it binds and transfers phosphatidylinositol and phosphatidylcholine between membrane compartments in vitro. Here we have used limited proteolysis of PITPα by subtilisin to identify the structural requirements for function. Digestion by subtilisin results in the generation of a number of slightly smaller peptide fragments, the major fragment being identified as a 29 kDa protein. The fragments were resolved by size-exclusion chromatography and were found to be totally inactive in both in vivo PLC reconstitution assays and in vitro phosphatidylinositol transfer assays. N-terminal sequencing and MS of the major 29 kDa fragment shows that cleavage occurs at the C-terminus of PITP at Met246, leading to a deletion of 24 amino acid residues. We conclude that the C-terminus plays an important role in mediating PLC signalling in vivo and lipid transfer in vitro, supporting the notion that lipid transfer may be a facet of PITP function in vivo.

An essential role for phosphatidylinositol transfer protein in phospholipase C-Mediated inositol lipid signaling

Cell ◽  
1993 ◽  
Vol 74 (5) ◽  
pp. 919-928 ◽  
Author(s):  
Geraint M.H. Thomas ◽  
Emer Cunningham ◽  
Amanda Fensome ◽  
Andrew Ball ◽  
Nicholas F. Totty ◽  
...  
Keyword(s):  
Phospholipase C ◽  
Essential Role ◽  
Lipid Signaling ◽  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

scholarly journals Phosphatidylinositol Transfer Protein Expression Altered by Aging and Parkinson Disease

2006 ◽  
Vol 26 (7-8) ◽  
pp. 1151-1164 ◽  
Author(s):  
Małgorzata Chalimoniuk ◽  
Gerry T. Snoek ◽  
Agata Adamczyk ◽  
Andrzej Małecki ◽  
Joanna B. Strosznajder
Keyword(s):  
Protein Expression ◽  
Parkinson Disease ◽  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

X-ray analysis of crystals of rat phosphatidylinositol-transfer protein with bound phosphatidylcholine

1999 ◽  
Vol 55 (2) ◽  
pp. 522-524 ◽  
Author(s):  
Randall L. Oliver ◽  
Jacqueline M. Tremblay ◽  
George M. Helmkamp ◽  
Lynwood R. Yarbrough ◽  
Natalie W. Breakfield ◽  
...  
Keyword(s):  
Crystal Form ◽  
Unit Cell ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
Vapor Diffusion ◽  
Solvent Content ◽  
Cell Parameters ◽  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

Phosphatidylinositol-transfer protein (PITP) is a soluble, ubiquitously expressed, highly conserved protein encoded by two genes in humans, rodents and other mammals. A cDNA encoding the alpha isoform of the rat gene was expressed to high levels in Escherichia coli, the protein purified and the homogeneous protein used for crystallization studies. Crystals of rat PITP-α were obtained by vapor-diffusion techniques using the sitting-drop method. Crystals grow within two weeks by vapor-diffusion techniques in the presence of polyethylene glycol 4000. Both crystal forms pack in the monoclinic space group P21. Crystal form I has unit-cell parameters a = 44.75, b = 74.25, c = 48.32 Å and β = 114.14°. Unit-cell parameters for crystal form II are a = 47.86, b = 73.59, c = 80.49 Å and β = 98.54°. Crystal form I has a Vm of 2.295 Å3 Da−1 and an estimated solvent content of 46.4% with one molecule per asymmetric unit, while crystal form II has a Vm of 2.196 Å3 Da−1 and an estimated solvent content of 44.0%, assuming two molecules per asymmetric unit.

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