4WL2

Structure of penicillin V acylase from Pectobacterium atrosepticum

Summary for 4WL2

• Entry DOI: 10.2210/pdb4wl2/pdb
• Descriptor: Putative exported choloylglycine hydrolase (2 entities in total)
• Functional Keywords: ntn hydrolase, choloylglycine, penicillin v, periplasmic, hydrolase
• Biological source: Pectobacterium atrosepticum SCRI1043
• Total number of polymer chains: 8
• Total formula weight: 315802.31

Authors

Ramasamy, S.,Avinash, V.S.,Pundle, A.V.,Suresh, C.G. (deposition date: 2014-10-06, release date: 2015-10-07, Last modification date: 2024-10-16)

Primary citation

Avinash, V.S.,Panigrahi, P.,Chand, D.,Pundle, A.,Suresh, C.G.,Ramasamy, S.
Structural analysis of a penicillin V acylase from Pectobacterium atrosepticum confirms the importance of two Trp residues for activity and specificity
J.Struct.Biol., 193:85-94, 2016

PubMed Abstract: Penicillin V acylases (PVA) catalyze the deacylation of the beta-lactam antibiotic phenoxymethylpenicillin (Pen V). They are members of the Ntn hydrolase family and possess an N-terminal cysteine as the main catalytic nucleophile residue. They form the evolutionarily related cholylglycine hydrolase (CGH) group which includes bile salt hydrolases (BSH) responsible for bile deconjugation. Even though a few PVA and BSH structures have been reported, no structure of a functional PVA from Gram-negative bacteria is available. Here, we report the crystal structure of a highly active PVA from Gram-negative Pectobacterium atrosepticum (PaPVA) at 2.5Å resolution. Structural comparison with PVAs from Gram-positive bacteria revealed that PaPVA had a distinctive tetrameric structure and active site organization. In addition, mutagenesis of key active site residues and biochemical characterization of the resultant variants elucidated the role of these residues in substrate binding and catalysis. The importance of residue Trp23 and Trp87 side chains in binding and correct positioning of Pen V by PVAs was confirmed using mutagenesis and substrate docking with a 15ns molecular dynamics simulation. These results establish the unique nature of Gram-negative CGHs and necessitate further research about their substrate spectrum.

PubMed: 26707624
DOI: 10.1016/j.jsb.2015.12.008

Experimental method

X-RAY DIFFRACTION (2.5 Å)