Screening Of A High Chitin Deacetylase-Producing Strain

Chitin is a straight-chain polymer compound which is formed by theN-acetyl-D-glucosamine monomer connected by β-1,4glycosidic bonds. Chitinis one of the most abundant natural organic compounds in nature, and theirquantity is just behind quantity of cellulose. Chitosan is the N-deacetylated formof chitin,which is widely used in many areas such as food, medicine, lightindustry, printing and dying, environmental protection and agriculture. Chitindeacetylase can generat chitosan by the way of removal of the acetyl group in thechitin.The degree of deacetylation of chitin pretreats up to97%if CDA roleinchitin. Deacetylation processing was mild, specificity and less poluted byenzymatic which is worth to investigating and developmentment.Currently thestudy of CDA in domestic and foreign was mainy focusing on several kinds offungi.This work isolated a high CDA-producing actinomycetes which is breededafter collecting soil samples and the two-step screening,identified the stain andstudied the fermentation conditions and the resulting deacetylase enzymaticproperties. The study will provid sources of isolation and screening of chitindeacetylase high-producing strains and preparation for the biological chitindeacetylase source, even provided research bases to further improve the strainproducing the enzyme capacity and enzyme activity.The experimental results are as follows:(1)A bacterium which produces a high activity chitin deacetylase has beenisolated from soil by color reaction and named as F2-7-3to screen the high chitindeacetylase-producing strains, provide sources of chitin deacetylase biologicalpreparation and the basis for further enhancing the ability of enzyme productionof the strains and enzyme activity. The strain was identified as Rhodococcus bymorphological, physiological and biochemical characteristics and16S rDNAanalysis. The enzymatic properties of chitin deacetylase which produced by fermentation of strains were studied. The strains initially identified that strainF2-7-3may be members of Actinomycetes door (Actinobacteria), actinomycetesGang (Actinobacteria), the actinomycetes subclass (Actinobacteridae),Actinomycetales (Actinomycetales), Corynebacterium suborder(Corynebacterineae), Nocardia Branch (Nocardiaceae), Rhodococcus genus(Rhodococcus).(2)By using single factor experiment and response surface analysis，fermentation medium composition and culture conditions has been studied，according to the preliminary studies, optimal culture conditions of chitindeacetylase production of the strain F2-7-3is30℃,pH7, the seed age15to24hand the10%liquid volume; the key factors of CDA production mediumcomposition has been selected: sucrose, yeast extract, KH2PO4,MgSO4, And theuse of the test fitting of Box-Behnken response surface function, responsesurface function has been work out with the test fitting of Box-Behnken. Activitylevel is260.26/mL in this condition.(3) The study of chitin deacetylase enzymatic properties which the strainF2-7-3produced shows that enzyme activity of the CDA will higher underneutral to slightly alkaline environment, the optimum pH was7.0, optimumtemperature was50℃, Mg2+,Zn2+,Fe2+and Co2+show inhibition to the enzymaticreaction, and the stronger the inhibition will increasing with the concentrationsof Metal ions. Ca2+,Mn2+and K+had activation effet, however, as theconcentration increases, showing a strong inhibitory effect. The analysis resultsshowed that the bacteria enzyme production ability, with good value fordevelopment.