Conformation and Lipid Binding Properties of Four Peptides Derived from the Membrane-Binding Domain of CTP:Phosphocholine Cytidylyltransferase†

Biochemistry ◽  
1998 ◽  
Vol 37 (26) ◽  
pp. 9509-9519 ◽  
Author(s):  
Joanne E. Johnson ◽  
N. Madhusudhana Rao ◽  
Sek-Wen Hui ◽  
Rosemary B. Cornell
Keyword(s):  
Membrane Binding ◽  
Lipid Binding ◽  
Binding Domain ◽  
Binding Properties ◽  
Ctp:Phosphocholine Cytidylyltransferase

scholarly journals Identification of an 11-residue portion of CTP–phosphocholine cytidylyltransferase that is required for enzyme–membrane interactions

1997 ◽  
Vol 325 (1) ◽  
pp. 29-38 ◽  
Author(s):  
Jilin YANG ◽  
Jinxia WANG ◽  
Irene TSEU ◽  
Maciej KULISZEWSKI ◽  
Wensu LEE ◽  
...  
Keyword(s):  
Oleic Acid ◽  
Fusion Proteins ◽  
Membrane Binding ◽  
Lipid Vesicles ◽  
Lipid Binding ◽  
Binding Domain ◽  
Membrane Interactions ◽  
Phosphocholine Cytidylyltransferase ◽  
N Terminus ◽  
L2 Cells

CTP–phosphocholine cytidylyltransferase (CT) is a key regulatory enzyme in the biosynthesis of phosphatidylcholine (PC) in many cells. Enzyme–membrane interactions appear to play an important role in CT activation. A putative membrane-binding domain appears to be located between residues 236 and 293 from the N-terminus. To map the membrane-binding domain more precisely, glutathione S-transferase fusion proteins were prepared that contained deletions of various domains in this putative lipid-binding region. The fusion proteins were assessed for their binding of [3H]PC/oleic acid vesicles. Fusion proteins encompassing residues 267–277 bound to PC/oleic acid vesicles, whereas fragments lacking this region exhibited no specific binding to the lipid vesicles. The membrane-binding characteristics of the CT fusion proteins were also examined using intact lung microsomes. Only fragments encompassing residues 267–277 competed with full-length 125I-labelled CT, expressed in recombinant Sf9 insect cells, for microsomal membrane binding. To investigate the role of this region in PC biosynthesis, A549 and L2 cells were transfected with cDNA for CT mutants under the control of a glucocorticoid-inducible long terminal repeat (LTR) promoter. Induction of CT mutants containing residues 267–277 in transfectants resulted in reduced PC synthesis. The decrease in PC synthesis was accompanied by a shift in endogenous CT activity from the particulate to the soluble fraction. Expression of CT mutants lacking this region in A549 and L2 cells did not affect PC formation and subcellular distribution of CT activity. These results suggest that the CT region located between residues 267 and 277 from the N-terminus is required for the interaction of CT with membranes.

scholarly journals The Ca2+-dependent Lipid Binding Domain of P120GAPMediates Protein-Protein Interactions with Ca2+-dependent Membrane-binding Proteinss

1996 ◽  
Vol 271 (40) ◽  
pp. 24333-24336 ◽  
Author(s):  
Alison J. Davis ◽  
Jonathan T. Butt ◽  
John H. Walker ◽  
Stephen E. Moss ◽  
Debra J. Gawler
Keyword(s):  
Protein Interactions ◽  
Membrane Binding ◽  
Lipid Binding ◽  
Binding Domain ◽  
Protein Protein Interactions

Structure of the Membrane Binding Domain of CTP:Phosphocholine Cytidylyltransferase†,‡

Biochemistry ◽  
1996 ◽  
Vol 35 (37) ◽  
pp. 11975-11984 ◽  
Author(s):  
Simon J. Dunne ◽  
Rosemary B. Cornell ◽  
Joanne E. Johnson ◽  
Nicholas R. Glover ◽  
Alan S. Tracey
Keyword(s):  
Membrane Binding ◽  
Binding Domain ◽  
Ctp:Phosphocholine Cytidylyltransferase

Amino acid substitutions in the membrane-binding domain of cytochrome b5 alter its membrane-binding properties

1993 ◽  
Vol 1153 (2) ◽  
pp. 163-169 ◽  
Author(s):  
Vera G. Tretyachenko-Ladokhina ◽  
Alexey S. Ladokhin ◽  
Liming Wang ◽  
Alan W. Steggles ◽  
Peter W. Holloway
Keyword(s):  
Amino Acid ◽  
Cytochrome B5 ◽  
Membrane Binding ◽  
Binding Domain ◽  
Amino Acid Substitutions ◽  
Binding Properties

scholarly journals Characterization of the lipid-binding domain of the Plasmodium falciparum CTP:phosphocholine cytidylyltransferase through synthetic-peptide studies

2003 ◽  
Vol 375 (3) ◽  
pp. 653-661 ◽  
Author(s):  
Marie-Pierre LARVOR ◽  
Rachel CERDAN ◽  
Catherine GUMILA ◽  
Luc MAURIN ◽  
Patrick SETA ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Surface Pressure ◽  
Hydrophobic Interactions ◽  
Lipid Binding ◽  
Binding Domain ◽  
Helical Conformation ◽  
Initial Surface ◽  
Water Interface ◽  
Ctp:Phosphocholine Cytidylyltransferase ◽  
Air Water Interface

Phospholipid biosynthesis plays a key role in malarial infection and is regulated by CCT (CTP:phosphocholine cytidylyltransferase). This enzyme belongs to the group of amphitropic proteins which are regulated by reversible membrane interaction. To assess the role of the putative membrane-binding domain of Plasmodium falciparum CCT (PfCCT), we synthesized three peptides, K21, V20 and K54 corresponding to residues 274–294, 308–327 and 274–327 of PfCCT respectively. Conformational behaviour of the peptides, their ability to bind to liposomes and to destabilize lipid bilayers, and their insertion properties were investigated by different biophysical techniques. The intercalation mechanisms of the peptides were refined further by using surface-pressure measurements on various monolayers at the air/water interface. In the present study, we show that the three studied peptides are able to bind to anionic and neutral phospholipids, and that they present an α-helical conformation upon lipid binding. Peptides V20 and the full-length K54 intercalate their hydrophobic parts into an anionic bilayer and, to a lesser extent, a neutral one for V20. Peptide K21 interacts only superficially with both types of phospholipid vesicles. Adsorption experiments performed at the air/water interface revealed that peptide K54 is strongly surface-active in the absence of lipid. Peptide V20 presents an atypical behaviour in the presence of phosphatidylserine. Whatever the initial surface pressure of a phosphatidylserine film, peptide V20 and phosphatidylserine entities seem linked together in a special organization involving electrostatic and hydrophobic interactions. We showed that PfCCT presents different lipid-dependence properties from other studied CCTs. Although the lipid-binding domain seems to be located in the C-terminal region of the enzyme, as with the mammalian counterpart, the membrane anchorage, which plays a key role in the enzyme regulation, is driven by two α-helices, which behave differently from one another.

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