6576

6141

كارشناسي ارشد

بيوتكنولوژي

اصفهان: دانشگاه صنعتي اصفهان، دانشكده كشاورزي

1390

پانزده، 142ص.: مصور، جدول، نمودار

ص.ع. به فارسي و انگليسي

آقافخرميرلوحي، علي اصغر كارخانه

فاطمه تابنده

21/1/91

آذر شاه پيري، محمدعلي ادريس

1396/10/10

كتابنامه

كشاورزي

مهندسي كشاورزي

ID6141

به فارسي و انگليسي: قابل ويت در نسخه ديجيتالي

The Effect of 5 Domain Deletion in Bacillus thermocatenulatusLipase on Enzyme s Properties Nastaran Goodarzi nastaran goudarzi@yahoo com Date of Submission Februery 7 2012 Department of Agricultural Biotechnology Isfahan University of Technology Isfahan 84156 83111 Iran Degree M Sc Language Farsi 1 A Mirlohi Ph D Professor Supervisor mirlohi@cc iut ac ir 2 A A Karkhane Ph D Assist Professor Supervisor karkhane@nigeb ac irAbstractLipases triacylglycerol acyl hydrolase EC 3 1 1 3 are the most important industrial enzymes that are used invarious industries including food detergents pharmaceutical products leather textile cosmetic and paper Bacteriallipases are member of hydrolase family that hydrolyz triacylglycerols at the water lipid interface Thecomparision of Bacillus lipases with hydrolase structure showed that 5 helix enter in Bacillus lipases duringthe evolution Improvement of catalytic activity and substrate specificity by using protein engineering is significantin commercial applications In this study after bioinformatic researches 5 helix from btl2 gene of Bacillusthermocatenulatus was deleted by SOE PCR method and mutant gene was cloned into TA vector Then wastransformed into E coli bacteria After the confirmation of cloning mutant gene was cloned into pPICZ Bexpression vector afterwards it was transformed to Pichia pastoris yeast Moreover after the confirmation ofexpression enzyme s activity of lipase purified with DE52 resin was measured with pH stat device The effect ofvarious substrate temperature thermostability pH metal ions detergents and organic solvents on normal andmutant enzymes was examined The results indicated that the mutant lipase than the native lipasein a wider range of pH The mutation has dramatically increased enzymemainly C8 substrate Mutant lipase tolerates 45 60detergents food and oleochemical industries The results demonstrated that the mutant enzymesin presence of different organic solvents detergents and some ions in comparison with native enzyme In terms ofthermostability mutant enzyme did not change perceptibly comparing to the native one Probably the deletion of 5helix causes less exposure of hydrophobic molecules to the surface and in some cases this turn may result theincreasing of enzyme activity Furthermore it is supposed that the mutation changed the active site of mutantenzyme and provided the ability of hydrolyzing substrate with longer hydrocarbon chain Generally the resultproves that the structure of 5 helix which belongs to Bacillus thermocatenulatus lipase is not necessary and thedeletion of it did not change the folding of enzyme In addition this deletion in some cases increased the activity ofenzyme which may lead to more application of this enzyme in different industries Keywords Bacillus themocatenulatus lipase hydrolase Pichia pastoris 5 helix proteinengineering

آقافخرميرلوحي، علي اصغر كارخانه

فاطمه تابنده

آذر شاه پيري، محمدعلي ادريس