深海热液区化能自养菌Caminibacter profundus氢酶及其对H_2的响应特点

Hydrogenase of deep-sea hydrothermal chemoautotrophic Caminibacter profundus and its response to H_2

化能自养菌中的氢酶在深海热液区生态系统的物质和能量转化中具有重要作用。以Caminibacter profundus为研究对象,通过设计PCR引物,克隆编码膜结合的类型I NiFe-氢酶大亚基基因序列hynL并进行生物信息学分析;研究hynL相对表达、甲基紫晶(MV)还原氢酶活性以及菌株生长对H2浓度变化的响应特点。结果表明,从C.profundus克隆获得864 bp的hynL基因片段,其编码的氨基端序列与Lebetimonas acidiphila的相似性为99%,与热液区化能自养的Epsilonproteobacteria D类群的类型I NiFe氢酶大亚基属同一分支。hynL相对表达量和MV还原的氢酶活性分别于12 h和24 h达到最高,此时菌体处于指数生长期;hynL相对表达量和MV还原的氢酶活性和菌株生长的最适H2浓度均为60%。提示C.profundus通过调控hynL的表达,响应环境中H2浓度的变化,以影响菌株能量代谢的催化过程和生长繁殖。

Hydrogenases of chemolithoautotrophic bacteria play important roles in material and energy transfer in deep-sea hydrothermal ecosystem.By designing PCR primers,a hynL gene encoding large subunit of membrane-bound group I NiFe hydrogenase from Caminibacter profundus was cloned and bioinformatively analyzed.The relative expressions of hynL,methyl viologen（MV）-reducing hydro-genase activities and bacterial growth in response to different H2 concentration were also studied.The results showed that a 864 bp hynL gene segment was obtained,predicted amino acid sequence had a 99% similarity with that of Lebetimonas acidiphila,and it belongs to the same phylogenetic branch with group I NiFe hydrogenase large subunit of hydrothermal chemolithoautotrophic Epsilonproteobacteria D group.Both hynL relative expression and MV-reducing hydrogenase activity reached the highest level at 12 h and 24 h respectively,when the strain was in the exponential growth phase.Sixty per cent of H2 concentration was optimal for hynL expression,MV-reducing hydrogenase activity and bacterial growth.All these results suggested that C.profundus regulates hynL expression in response to H2 changes in hydrothermal environment,thereby affecting the catalytic energy process,as well as the growth and propagation.