[en] The BlaB metallo-beta-lactamase of Chryseobacterium meningosepticum CCUG4310 was overproduced in Escherichia coli by means of a T7 promoter-based expression system. The overproducing system, scaled up in a 15-liter fermentor, yielded approximately 10 mg of BlaB protein per liter, mostly released in the culture supernatant. The enzyme was purified by two ion-exchange chromatographic steps with an overall yield of 66%. Analysis of the kinetic parameters revealed efficient activities (k(cat)/K-m ratios of >10(6) M-1 s(-1)) toward most penam and carbapenem compounds, with the exception of the 6-alpha-methoxypenam derivative temocillin and of biapenem, which were poorer substrates. Hydrolysis of cephalosporins was overall less efficient, with a remarkable variability that was largely due to variable affinities of the BlaB enzyme for different compounds. BlaB was also able to hydrolyze serine-beta-lactamase inhibitors, including beta-iodopenicillanate, sulbactam and, although less efficiently, tazobactam.
Disciplines :
Pharmacy, pharmacology & toxicology Microbiology
Author, co-author :
Vessillier, S.
Docquier, Jean-Denis ; Université de Liège - ULiège > Département des sciences de la vie > Centre d'ingénierie des protéines
Rival, S.
Frère, Jean-Marie ; Université de Liège - ULiège > Département des sciences de la vie > Département des sciences de la vie
Galleni, Moreno ; Université de Liège - ULiège > Département des sciences de la vie > Macromolécules biologiques
Amicosante, G.
Rossolini, G. M.
Franceschini, N.
Language :
English
Title :
Overproduction and biochemical characterization of the Chryseobacterium meningosepticum BlaB metallo-beta-lactamase
Publication date :
June 2002
Journal title :
Antimicrobial Agents and Chemotherapy
ISSN :
0066-4804
eISSN :
1098-6596
Publisher :
Amer Soc Microbiology, Washington, United States - Washington
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