The Construction Of Fatty Acid Analysis Enzyme Thermal Sensor And The Purifity Of Trichoplusia Ni GV ENHANCIN

Diabetes is a worldwide disease,which etiology and pathogenesis is complex,so that they has not been fully elucidated so far,but the two main reasons is caused by f insulin resistance and the deficiency of islet beta cells secretion two factors.It is now accepted that FFA has played an important role in the occurrence of these two factors.Long-chain fatty acyl coenzyme A synthetase can detect FFA in vitro,which function is to catalyze oleic acid born long chain fatty acids combining with coenzyme A to obtain long-chain fatty acyl coenzyme A,our research object is the long-chain fatty acyl coenzyme A synthetase FadD1 of the pseudomonas aeruginosa PAOl.the following is our mainly research:1)Using PAOl genome as templates,amplification of fadDl gene is finished.Sequence analysis found that fadDl open reading frame(open reading frame,ORF)is 1689 bp,encoding 562 amino acid residues,whose molecular weight is about 61.7 kDa.Structure domain analysis showed that fadD1 contains 1 AMP binding functional domains.fadDl gene was cloned into a cloning vector,then we construct the recombinant expression vector,achieving the expression of recombinant proteins in e.coli.2)After purifing FadD1 and detecting enzyme activity,purified product of FadDl was fixed to the aperture controllable glass ball,building the enzyme thermal sensors and playing a preliminary test.Our research object,pseudomonas aeruginosa long-chain fatty acyl coenzyme A synthetase FadDl,can test FFA in vitro.FadD1 fixed on enzyme thermal sensor can detect heat produced by enzymatic reaction,analyzing the content of free fatty acids,which will make FFA sensitive detection more rapid,stable and reliable,so it provides a new method for the early diagnosis of diabetes.In addition,Additional work we do is that we purified Enhancin of Trichoplusia ni granulosis virus(TnGV)and carried incubation with the cotton boll worm peritrophic membrane in vitro in order to obtain peritrophic membrane targets.