Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Characterization of Glutaryl 7-ACA Acylase from Pseudomonas diminuta KAC-1

  • Kim, Dae-Weon (School of Food Biotechnology and Nutrition, Woosong University) ;
  • Kang, Sang-Mo (Department of Microbiological Engineering, Kon-kuk Unviersity) ;
  • Yoon, Ki-Hong (School of Food Biotechnology and Nutrition, Woosong University)
  • Published : 2001.06.01
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Abstract

The glutaryl 7-aminocephalosporanic acid (glutaryl 7-ACA) acylase was purified from Pseudomonas diminuta KAC-1 cells isolated from soil, and characterized. The acylase was purified by procedures including ammonium sulfate fractionation and column chromatographies on DEAE-Sepharose, Phenyl-Sepharose, Q-Sepharose, and Superose 12H/R. The negative acylase was found to be composed of two subunits with molecular masses of approximately 55 kDa and 17 kDa, respectively. The isoelectric point of the enzyme was 4.0. The specific activities of the purified acylase were 8.0 and 7.0 U/mg on glutaryl 7-ACA and glutaryl 7-aminodesacetoxy cephalosporanic acid (glutaryl 7-ADCA), respectively, and $K_m$ values were 0.45 mM for glutaryl 7-ADCA and 0.67 mM for glutaryl 7-ADCA. The enzyme had a pH optimum at 8.0 and a tmperature optimum at $40^{\circ}C$. The acylase catalyzed the synthesis of glutaryl 7-ACA from glutaric acid and 7-ACA as well as the hydrolysis of glutaryl 7-ADCA, although the reaction rate of the synthesis was slower than that of the hydrolysis. In addition, it was found that the enzyme had a glutaryl transferase activity, thereby transferring the glutaryl group from one cephalosporin nucleus to another.

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