title: Suggestive evidence for the involvement of the second calcium and surface loop in interfacial binding: monoclinic and trigonal crystal structures of a quadruple mutant of phospholipase A2.
abstract: The crystal structures of the monoclinic and trigonal forms of the quadruple mutant K53,56,120,121M of recombinant bovine pancreatic phospholipase A2 (PLA2) have been solved and refined at 1.9 and 1.1 A resolution, respectively. Interestingly, the monoclinic form reveals the presence of the second calcium ion. Furthermore, the surface-loop residues are ordered and the conformation of residues 62-66 is similar to that observed in other structures containing the second calcium ion. On the other hand, in the trigonal form the surface loop is disordered and the second calcium is absent. Docking studies suggest that the second calcium and residues Lys62 and Asp66 from the surface loop could be involved in the interaction with the polar head group of the membrane phospholipid. It is hypothesized that the two structures of the quadruple mutant, monoclinic and trigonal, represent the conformations of PLA2 at the lipid interface and in solution, respectively. A docked structure with a phospholipid molecule and with a transition-state analogue bound, one at the active site coordinating to the catalytic calcium and the other at the second calcium site, but both at the i-face, is presented.
authors.affiliation: Bioinformatics Centre, Indian Institute of Science, Bangalore 560 012, India. sekar@physics.iisc.ernet.in
authors.forename: K
authors.initials: K
authors.lastname: Sekar
authors.forename: M
authors.initials: M
authors.lastname: Yogavel
authors.forename: Shankar Prasad
authors.initials: SP
authors.lastname: Kanaujia
authors.forename: Alok
authors.initials: A
authors.lastname: Sharma
authors.forename: D
authors.initials: D
authors.lastname: Velmurugan
authors.forename: M-J
authors.initials: MJ
authors.lastname: Poi
authors.forename: Z
authors.initials: Z
authors.lastname: Dauter
authors.forename: M-D
authors.initials: MD
authors.lastname: Tsai
mesh_terms: D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry*; D002417:Cattle; D003198:Computer Simulation; D003460:Crystallization / Q000379:methods; D018360:Crystallography, X-Ray; D054513:Group IV Phospholipases A2; D006860:Hydrogen Bonding; D006868:Hydrolysis; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry*; D020125:Mutation, Missense; D010179:Pancreas / Q000201:enzymology; D010741:Phospholipases A / Q000737:chemistry* / Q000235:genetics; D054467:Phospholipases A2; D010743:Phospholipids; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D013552:Swine
chemical_list: D050505:Mutant Proteins; D010743:Phospholipids; D011994:Recombinant Proteins; D010741:Phospholipases A; D054513:Group IV Phospholipases A2; D054467:Phospholipases A2; D002118:Calcium