Comparison between apo and complexed structures of bothropstoxin-I reveals the role of Lys122 and Ca(2+)-binding loop region for the catalytically inactive Lys49-PLA(2)s.
Phospholipases A(2) (Asp49-PLA(2)s) are enzymes responsible for cellular membrane disruption through Ca(2+)-dependent hydrolysis of phospholipids. A class of these proteins (Lys49-PLA(2)s) does not show catalytic activity but can exert a pronounced local myotoxic effect that is not neutralized by serum therapy. In this work, we present five structures of Lys49-PLA(2)s from snakes of the Bothrops genus in apo form, complexed with PEG molecules and chemically modified by p-bromofenacil bromide (BPB), a classic inhibitor of PLA(2). We present herein an extensive structural analysis including: (i) the function of hydrophobic long-chain molecules as Lys49-PLA(2)s inhibitors, (ii) the role of Lys122, previously indicated as being responsible for Lys49-PLA(2)s catalytic inactivity and, (iii) a structural comparison of the Ca(2+)-binding loop region between Lys49 and Asp49-PLA(2)s. The Lys122 analysis of 30 different monomers for apo and complexed Lys49-PLA(2)s structures shows that this residue is very flexible and may bind to different carboxyl groups giving stability to the crystal structures. The structural comparisons of the Ca(2+)-binding loop region between Lys49 and Asp49-PLA(2)s reveal the importance of the Tyr28 residue conservation in Asp49-PLA(2)s to the integrity of this loop. The Tyr28 residue stabilizes this region by an interaction with Gly35 residue. In Lys49-PLA(2)s and low-catalytic Asp49-PLA(2)s this interaction does not occur, preventing the binding of Ca(2+).
Fernandes
Carlos A H
CA
Departamento de Física e Biofísica, Instituto de Biociências, UNESP - Univ Estadual Paulista, Botucatu, SP, Brazil.
Marchi-Salvador
Daniela P
DP
Salvador
Guilherme M
GM
Silva
Mabel C O
MC
Costa
Tássia R
TR
Soares
Andreimar M
AM
Fontes
Marcos R M
MR
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D017837:Bothrops; D002118:Calcium / Q000737:chemistry* / Q000502:physiology; D003435:Crotalid Venoms / Q000737:chemistry*; D018360:Crystallography, X-Ray; D008239:Lysine / Q000737:chemistry* / Q000502:physiology; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D013329:Structure-Activity Relationship
D003435:Crotalid Venoms; C078097:bothropstoxin; D008239:Lysine; D002118:Calcium