Kesado T, Lindqvist L, Hedberg M, Tunér K, Nord C E
Department of Microbiology, Huddinge University Hospital, Karolinska Institute, Sweden.
Antimicrob Agents Chemother. 1989 Aug;33(8):1302-7. doi: 10.1128/AAC.33.8.1302.
A highly penicillin-resistant beta-lactamase-producing strain of Clostridium butyricum, strain NBL 3, was isolated. The specific activity of the unpurified beta-lactamase was 0.29 U/mg of protein. The enzyme was purified 1,130-fold by QAE Zetaprep, Sephacryl S-300, and Mono Q column passages. The purified enzyme was judged homogeneous by sodium dodecyl sulfate gradient gel electrophoresis and fast protein liquid chromatography. The enzyme hydrolyzed phenoxymethylpenicillin, benzylpenicillin, ampicillin, and carbenicillin more rapidly than piperacillin and cephaloridine. Cephalothin, cephalexin, cefoxitin, moxalactam, cefotaxime, and imipenem were only slightly hydrolyzed, at less than 1% the rate for phenoxymethylpenicillin. The enzyme was inhibited by clavulanic acid, sulbactam, tazobactam, and p-chloromercuribenzoate. The molecular weight was determined by gel filtration and sodium dodecyl sulfate gradient gel electrophoresis to be 32,000. The isoelectric point was 4.4. Aspartic acid-asparagine, glutamic acid-glutamine, leucine, lysine, alanine, and serine dominated the amino acid composition.
分离出了一株丁酸梭菌的高产青霉素耐药性且产生β-内酰胺酶的菌株NBL 3。未纯化的β-内酰胺酶的比活性为0.29 U/mg蛋白质。通过QAE Zetaprep、Sephacryl S-300和Mono Q柱层析，该酶被纯化了1130倍。通过十二烷基硫酸钠梯度凝胶电泳和快速蛋白质液相色谱法判断纯化后的酶为均一的。该酶水解苯氧甲基青霉素、苄青霉素、氨苄青霉素和羧苄青霉素的速度比哌拉西林和头孢菌素更快。头孢噻吩、头孢氨苄、头孢西丁、莫拉酰胺、头孢噻肟和亚胺培南仅被轻微水解，水解速率不到苯氧甲基青霉素的1%。该酶受到克拉维酸、舒巴坦、他唑巴坦和对氯汞苯甲酸的抑制。通过凝胶过滤和十二烷基硫酸钠梯度凝胶电泳测定分子量为32000。等电点为4.4。天冬氨酸-天冬酰胺、谷氨酸-谷氨酰胺、亮氨酸、赖氨酸、丙氨酸和丝氨酸在氨基酸组成中占主导地位。
