TAILIEUCHUNG - Báo cáo khoa học: Determination of the metal ion dependence and substrate specificity of a hydratase involved in the degradation pathway of biphenyl/chlorobiphenyl

BphH is a divalent metal ion-dependent hydratase that catalyzes the forma-tion of 2-keto-4-hydroxypentanoate from 2-hydroxypent-2,4-dienoate (HPDA). This reaction lies on the catabolic pathway of numerous aromat-ics, including the significant environmental pollutant, polychlorinated biphenyls (PCBs). BphH from the PCB degrading bacterium,Burkholderia xenoveransLB400, was overexpressed and purified to homogeneity. | iFEBS Journal Determination of the metal ion dependence and substrate specificity of a hydratase involved in the degradation pathway of biphenyl chlorobiphenyl Pan Wang and Stephen Y. K. Seah Department of Microbiology University of Guelph Ontario Canada Keywords aromatics hydratase 2-hydroxypent-2 4-dienoate metal cofactor substrate specificity Correspondence S. Y. K. Seah Department of Microbiology University of Guelph Guelph Ontario Canada N1G 2W1 Fax 1 519 837 1802 Tel 1 519 824 4120 Ext. 56750 E-mail sseah@ Received 22 October 2004 revised 9 December 2004 accepted 15 December 2004 doi BphH is a divalent metal ion-dependent hydratase that catalyzes the formation of 2-keto-4-hydroxypentanoate from 2-hydroxypent-2 4-dienoate HPDA . This reaction lies on the catabolic pathway of numerous aromatics including the significant environmental pollutant polychlorinated biphenyls PCBs . BphH from the PCB degrading bacterium Burkholderia xenoverans LB400 was overexpressed and purified to homogeneity. Atomic absorption spectroscopy and Scatchard analysis reveal that only one divalent metal ion is bound to each enzyme subunit. The enzyme exhibits the highest activity when Mg2 was used as cofactor. Other divalent cations activate the enzyme in the following order of effectiveness Mg2 Mn2 Co2 Zn2 Ca2 . This differs from the metal activation profile of the homologous hydratase MhpD. UV-visible spectroscopy of the Co2 -BphH complex indicates that the divalent metal ion is hexa-coordinated in the enzyme. The nature of the metal ion affected only the kcat and not the Km values in the BphH hydration of HPDA suggesting that cation has a catalytic rather than just a substrate binding role. BphH is able to transform alternative substrates substituted with methyl-and chlorine groups at the 5-position of HPDA. The specificity constants kcat Km for 5-methyl and 5-chloro substrates are however lowered by eight- and 67-fold compared with the .

• Báo cáo khoa học
• Report medicine
• traditional medicine
• scientific reports
• biochemical studies
• environmental medicine
• Crystal structure
• medical knowledge
• medical research
• analysis of cytoplasmic
• oxidation
• bacteria
• hemoglobin
• medicine
• cell proliferation
• breast cancer
• chemical reaction
• gastric cancer
• hormone medicine
• tumor cells