肺炎克雷伯杆菌β-内酰胺酶的分子特性

目的 探讨头孢哌酮（MIC≥8mg/L）加舒巴坦后头孢哌酮的最低抑菌浓度（MIC）降低2倍以上的肺炎克雷伯杆菌β－内酰胺酶分子特性。方法 用PCR扩增和DNA测序方法,测定分子量及等电点以及酶动力。结果 4株为克雷伯杆菌含有TEM型基因,2株肺炎克雷伯杆菌TEM型酶的氨基酸序列与TEM－1相比在117位及118位发生了氨基酸变异。4株肺炎克雷伯杆菌都产生2种以上的酶,等电点从5.4-9.3,相对分子量从23000－34000。酶动力学研究表明,肺炎克雷伯杆菌中99592及99607产生的酶能够水解头孢他啶、头孢噻肟及头孢曲松,并且99592的酶活性大于99607的酶活性。结论 肺炎克雷伯杆菌可能产生新的抑制剂敏感的TEM型超广谱酶。

Molecular characteristics of beta-lactamase from Klebsiella pneumoniae

OBJECTIVE: To study the molecular characteristics of beta-lactamase from Klebsiella pneumoniae. METHODS: Beta-lactamase was prepared from strains of Klebsiella pneumoniae isolated clinically for which the minimal inhibitory concentration (MIC) of cefoperazone had been >or= 8 mg/L and then was reduced by 50% by the addition of sulbatam. The isoelectric points of different beta -lactamases were examined by LKB2117 Multiphor II Electrophoresis System. The molecular weights were examined by polyacrylamide gel electrophoresis. PCR was used to detect the TEM-type genes. The PCR products were sequenced by chemiluminescence. RESULTS: All four strains of Klebsiella pneumoniae produced more than two kinds of beta-lactamase. TEM-type genes were amplified from all strains. 97% - 98% of the DNA sequence and amino acid sequence of the TEM-type enzymes from 2 strains of Klebsiella pneumoniae were identical with those of TEM-1 enzyme. The isoelectric points of the beta-lactamases ranged from 5.4 to 9.30 and the molecular weights were between 23.0 and 43.0 KD. The beta-lactamases from Klebsiella pneumoniae 99592 and K. pneumoniae 99607 hydrolysed ceftazidine, cefotaxime, and ceftriaxone. The beta-lactamase from K. pneumoniae 99595 had stronger activity against ceftazidime, ceftaxime, and ceftriaxone than that from K. pneumoniae 99607. CONCLUSION: Klebsiella pneumoniae produces TEM-type extended spectrum beta -lactamase against ceftazidine, ceftaxime, and ceftriaxone.