Screening Of Cellulase Production Marine Strain And Studies On Enzyme-producing Conditions And Charaters Of Enzyme

Cellulose is the most abundant renewable natural biological resocurce on the earth,the biofuels from cellulosic materials is great significance for the sustainable development of human beings.Cellulase can convert cellulose into glucose which is the starting materials of biofuels fermentation.There are many types of cellulase production species which is widely spread in microorganisms,animals and even plants.The application of cellulase has been extended to many domains,such as food industry,animal husbandry,textile and laundry industry,pulp and paper industry.The particularity of the marine environment prompts marine microorganisms to generate multifarious enzyme systems.Therefore,marine microbial enzymes can offer novel biocatalysts with extraordinary properties.The development and utilization of marine microorganisms' enzyme resources can greatly enrich microbial enzymes repository.One cellulase production actinomycete strain was isolated from the sample collected from area of Taohua Island Zhoushan City Zhejiang Provinces.The strain was identified as Streptomyces pactum by morphological,physiological and biochemical characteristics combined with 16S rRNA gene sequence analysis.The optimal culture medium and condition of fermentation were study.The results indicated that the optimal carbon source and nitrogen source were bran and NH4NO3 respectively.Then analyszed the significant of culture medium components,which indicated that bran,K2HPO4,tween-80 is the significant factors influencing the cellulase production.The result of response surface analysis of the three significant factors determined the best composition of culture medium.At the same time,the genetic algorithm with BP artificial neural network method was used to optimize the culture medium composition.Finally,optimize the culture condition based on the optimal culture medium.The optimal fermentation conditions as follows:fermentation temperature 30?,initial inoculum 5%,loading volume 70 mL/150 mL,shake flask speed 200 r/min,initial pH value 7,the of peak cellulase activity was in 168 hours 0.455 U/mL.Cellulase activity was increased by 97.82%.The crude enzymes were concentrated and prepared by ammonium sulfate precipitation and desalting treatment to obtain the preliminary purification cellulase.Properties of the preliminary purification cellulase were investigated.The result showed the optimal temperature was 60?,and the optimal pH between 6 and 7.The enzyme activity was improved by a certain concentration(?0.6%)NaCl,with 2%NaCl the enzyme activity was highest increase 1.19 fold than NaCl free.The enzyme was stabe below 50?.K+,Mg2+,Zn2+ could significantly promote enzyme activity,Fe2+,Cu2+,Co2+,Mn2+ could restrain.enzyme activity at low concentration(5mmol/L),and K+,Mg2+ could significantly promote enzyme activity,Fe2+,Cu2+,Co2+,Mn2+ could restrain enzyme activity at high concentration(10mmol/L).The properties of enzyme about salt tolerance,alkali tolerance and heat tolerance showed the potential in industrial applications.