| The histidine kinase-response regulator hybrid protein(HK)was identified as a member of two-component signaling systems in Gluconobacter oxydans based on domain organization analysis.Two-component signaling systems are wildly distributed in bacteria and plants,allowing these organisms to sense and respond to various environmental stimuli.Two-component signaling systems regulate a variety of essential processes such as cell-wall production,antibiotic resistance,osmoregulation,metabolism,transport,chemotaxis,pathogenicity,photosensitivity and so on.To clarify the potential target proteins of HK in cells,a yeast two-hybrid assay against G.oxydans 621H genome library with HK as a bait was conducted.The interactions between HK and its target proteins including histidine kinase,2-dehydro-3-deoxyglycolic acid aldolase and 1-deoxy-D-xylulose-5-phosphate synthase were further verified with GST pull-down assay and bimolecular fluorescence complementation assay in vitro and in vivo,respectively.Sequence analysis on three of those interactive proteins from G.oxydans showed thatGOX1641,containing a histidine kinase like ATPase domain without response regulator domain,is a putative histidine protein kinase,while GOX2281 encodes a 2-dehydro-3-deoxyglycolic acid aldolase which plays an important role in the growth of Gram-negative bacteria,and GOX0252 encodes an 1-deoxy-D-xylulose-5-phosphate synthase,also refered as DXS,which can be used for biosynthesis of thiamine(vitamin B1)and pyridoxine(vitamin B6).The gene sequence of DXS is highly conserved in those organisms.The determination of the interaction between the two-component protein HK and somemetabolism-related proteins in G.oxydans 621H indicated a possible role of HK in regulation on the metabolism in the bacterium in response to various enviromental stimuli,facilitating our understanding on the metabolic pathways and their regulation under stress. |
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