Identification of functionally important amino acids in a novel indigo-producing oxygenase from Rhodococcus sp. strain T104

Na Ra Kwon, Jong Chan Chae, Ki Young Choi, Miyoun Yoo, Gerben J. Zylstra, Young Min Kim, Beom Sik Kang, Eungbin Kim

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10 Citations (Scopus)

Abstract

A novel indigo-producing oxygenase gene, designated ipoA (1,197 bp) was characterized from Rhodococcus sp. strain T104. Three indigo-negative mutations (A58V, P59L, and G251D) were obtained through random mutagenesis using an E. coli mutator strain. Subsequent saturation mutagenesis resulted in the identification of nine and three amino acid substitutions that restore activity in the A58V and P59L mutants, respectively. Activity was not restored in the G251D mutation by any other amino acids. Interestingly, activity in the A58V mutant, where a methyl group is only replaced by an isopropyl side chain, is restored by a variety of amino acids, including polar ones. A molecular modeling study suggests that the residues at positions 58, 59, and 251 of the T104 IpoA enzyme are far from the active site, indicating that the mutations must alter the overall structure of the enzyme.

Original languageEnglish
Pages (from-to)417-422
Number of pages6
JournalApplied Microbiology and Biotechnology
Volume79
Issue number3
DOIs
Publication statusPublished - 2008 Jun

Bibliographical note

Funding Information:
Acknowledgment This work was supported by a grant from the Ministry of Science and Technology, Republic of Korea through the 21C Frontier Microbial Genomics and also in part by the Basic Research Program of the Korea Science and Engineering Foundation (Grant No. R01-2006-000-10136-0). NRK and KYC are recipients of Brain Korea 21 scholarships. GJZ acknowledges the support of NSF through grants MCB-0078465 and CHE-0221978.

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Applied Microbiology and Biotechnology

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