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Biochemistry (Moscow)

, Volume 73, Issue 1, pp 56–64 | Cite as

Molecular modeling studies of substrate binding by penicillin acylase

  • G. G. Chilov
  • O. V. Stroganov
  • V. K. ŠvedasEmail author
Article

Abstract

Molecular modeling has revealed intimate details of the mechanism of binding of natural substrate, penicillin G (PG), in the penicillin acylase active center and solved questions raised by analysis of available X-ray structures, mimicking Michaelis complex, which substantially differ in the binding pattern of the PG leaving group. Three MD trajectories were launched, starting from PDB complexes of the inactive mutant enzyme with PG (1FXV) and native penicillin acylase with sluggishly hydrolyzed substrate analog penicillin G sulfoxide (1GM9), or from the complex obtained by PG docking. All trajectories converged to a similar PG binding mode, which represented the near-to-attack conformation, consistent with chemical criteria of how reactive Michaelis complex should look. Simulated dynamic structure of the enzyme-substrate complex differed significantly from 1FXV, resembling rather 1GM9; however, additional contacts with residues bG385, bS386, and bN388 have been found, which were missing in X-ray structures. Combination of molecular docking and molecular dynamics also clarified the nature of extremely effective phenol binding in the hydrophobic pocket of penicillin acylase, which lacked proper explanation from crystallographic experiments. Alternative binding modes of phenol were probed, and corresponding trajectories converged to a single binding pattern characterized by a hydrogen bond between the phenol hydroxyl and the main chain oxygen of bS67, which was not evident from the crystal structure. Observation of the trajectory, in which phenol moved from its steady bound to pre-dissociation state, mapped the consequence of molecular events governing the conformational transitions in a coil region a143-a146 coupled to substrate binding and release of the reaction products. The current investigation provided information on dynamics of the conformational transitions accompanying substrate binding and significance of poorly structured and flexible regions in maintaining catalytic framework.

Key words

penicillin acylase substrate binding molecular modeling conformational flexibility 

Abbreviations

MD

molecular dynamics

PA

penicillin acylase

PG

penicillin G

PGSO

penicillin G sulfoxide

RHF

Restricted Hartree-Fock method

RMSD

root mean square deviation

RMSF

root mean square fluctuation

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Supplementary material

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Copyright information

© Pleiades Publishing, Ltd. 2008

Authors and Affiliations

  • G. G. Chilov
    • 1
  • O. V. Stroganov
    • 2
  • V. K. Švedas
    • 1
    • 2
    Email author
  1. 1.Belozersky Institute of Physico-Chemical BiologyLomonosov Moscow State UniversityMoscowRussia
  2. 2.Faculty of Bioengineering and BioinformaticsLomonosov Moscow State UniversityMoscowRussia

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