Study On The Screening And Enzymatic Properties Of A Noel Cold-adapted Amylase-producing Strain From Alpine Frozen Soils In The Tianshan Mountains

Object:Amylase is one of the most important enzyme for industry, which is widely used in several vocation like food industry, detergent industry, medicine industry et al. Cold-adapted amylase is having extensive application prospect in the directions of biological engineering and energy source saving because of its high catalytic activity and temperature sensitive. Generally, Cold-adapted amylase is product by the cold-adapted microorganisms, consequently, the screening of cold-adapted amylase-producing strains is the most important and fundamental factor to solve the source of Cold-adapted amylase. Frozen soil which called the resource-treasure house of cold-adapted microorganisms, is useful for us to study cold-adapted microorganisms and their cold active enzymes. Thus, isolation and purification of cold-adapted microorganisms from alpine frozen soils in the Tianshan Mountains is having great significance for screening the high-quality and high-activity cold-adapted enzymes-producing strains.Methods:This study depends on the distinct geographical environment of Xinjiang Province, isolating and purifying cold-adapted amylase-producing strains from Tianshan glacier, solving the source of Cold-adapted amylase. This study research on the enzymology property and the laboratory fermentation condition of cold-adapted enzymes-producing strains which are screening from alpine frozen soils in the Tianshan Mountains, to establish theory and practice foundation of elementary study and industrialization application of Cold-adapted amylaseResults:①Three strains producing cold-adapted amylase was isolated from frozen soil cont aining ice in Tianshan glacial by conventional deliquation plate process, which are sig ned L10-1,M12-1,X2. Physiological and biochemical identification of common bacteria and the phylogenetic development of strains indicated that:strain M12-land X2 are si-milar with Paenibacillus polymyxa(similarity:99%), strain L10-1 is similar with Hymen obacter.sp(similarity:96%) which indicated that strain L10-1 was temporary attached to Hymenobacter genus, waiting for further identification.②We determine the strain L10-1 as the objective strain of enzyme production by contrasting the optimum growth temperature,the optimization effective temperature of enzyme and the relative value of the average diameter and transparentphere, its relative enzyme activity has achieved 28.2U/ml.The optimum growth temperature section of this strain is 18～24℃, which belongs to psychrotrophs;the optimization effective temperature of amylase is 35℃, which is 25～30℃lower than the isozyme(the optimization effective temperature of the common temperature enzyme is 60～65℃)③Study on the enzymology property shows that the amylase producted by the strain L10-1 was an extracellular enzyme, and the optimization effective temperature of amylase is 35℃; the optimal pH value of enzyme is 6.0; it has a bad thermostable, which was stable temperature in 30℃and 40℃for half an hour which remained 60% and 20% of enzyme activity and almost has 0 enzyme activity when it was stable temperature in 60℃for 20 min; in addition,the amylase is stimulated in the presence of Ca2+,Mg2+,K+,Mn2+,Na+,but is inhibited by Co2+,Cu2+,Zn2+; phenylmethyl sulfonylfluoride(PMSF), ethylenediaminetetraacetic acid(EDTA), sodium dodecylsulfate(SDS), dimethyl sulphoxide (DMSO) showed partially inhibition on this enzyme, indicate that this enzyme was an metal activated enzyme and that a serine group was involved in catalytic center of this enzyme.④Study on the laboratory fermentation condition of cold-adapted enzymes-producing strains shows that The fermentation conditions were established as follow:The optimal carbon source of enzyme is sucrose and its optimal concentration is 1%; The optimal nitrogen source of enzyme is casein hydrolysate which optimal concentration is 1.5%; 500ml Erlenmeyer flask contains 250mL liquidness medium; the optimal rotation speed of shaking table was 200r/min; optimal fermentation time was 36h. Under these conditions the activity of cold-adapted amylase was improved from 15.23U/ml to 29.13U/ml.Conclusions:The conclusion all above is similar to the other's research which reported in China and abroad.Based on such characteristics, the enzyme should have a potential value in the industrial applications.