Crystallization and structure elucidation of GDSL esterase of Photobacterium sp. J15

Abstract

GDSL esterase J15 (EstJ15) is a member of Family II of lipolytic enzyme. The enzyme was further classified in subgroup SGNH hydrolase due to the presence of highly conserve motif, Ser-Gly-Asn-His in four conserved blocks I, II, III, and V, respectively. X-ray quality crystal of EstJ15 was obtained from optimized formulation containing 0.10 M ammonium sulphate, 0.15 M sodium cacodylate trihydrate pH 6.5, and 20% PEG 8000. The crystal structure of EstJ15 was solved at 1.38 Å with one molecule per asymmetric unit. The structure exhibits α/β hydrolase fold and shared low amino acid sequence identity of 23% with the passenger domain of the autotransporter EstA of Pseudomonas aeruginosa. The active site is located at the centre of the structure, formed a narrow tunnel that hinder long substrates to be catalysed which was proven by the protein-ligand docking analysis. This study facilitates the understanding of high substrate specificity of EstJ15 and provide insights on its catalytic mechanism.

Structure elucidation and docking analysis of 5M mutant of T1 lipase Geobacillus zalihae

“…The growth of disordered crystals was attributed to the uncontrolled and rapid nucleation of the crystal having overload protein concentration [29]. Similarly, Mazlan et al [30] verified that at greater than 10 mg/mL concentrations of protein, the quality of the crystal from GDSL esterase (EstJ15) of Photobacterium sp. was found not to be significantly improved.…”

Identification and Biochemical Characterization of a Novel Hormone-Sensitive Lipase Family Esterase Est19 from the Antarctic Bacterium Pseudomonas sp. E2-15

“…Because the side-chain of Glu had one more methylene group than that of Asp, the substrate pocket of E154D presented a deeper binding site compared to the wild Est19 ( Figure 5 ). This deeper site may make it more difficult for smaller substrates to orient toward catalytic residues [ 35 , 36 ], while, conversely, enabling substrates with longer acyl chains have better access to increase the relative activity toward p NPC10 and p NPC12 substrates. Furthermore, the E154D variant resulted in the loss of activity toward methyl L-lactate, indicating that Glu 154 may influence the enantioselectivity of Est19.…”