L-1,3 Glucannase 유전자의 Streptomyceslividans에서의 cloning 및 발현
Cloning and Expression of α-1,3 glucanase from Streptomyces sp. SW403 into Streptomyces lividans.
윤정원, 홍성현, 한홍근,
소속 상세정보
윤정원 ( ) - 수원대학교
홍성현 ( ) - 수원대학교
한홍근 ( ) - 수원대학교
KMID : 0355419950190040393
Abstract
Insoluble glucan produced by Streptococcus mutans has been implicated as causative agents of dental caries. Insoluble glucan produced by oral microorganisms is very hardly degradable by any dextranase because which contained α-1,3 glucosidic
bond
over
95%. Streptomyces sp. SW403 which produced α-1,3 glucanase was found from soil but it's productivity was low and unstable. The α-1,3 glucanase gene from Streptomyces sp. SW403 was cloned into Streptomyces lividans 1326JI with pIJ702. Genomic
DNA
from
Streptomtces sp. SW403 was partially digested with the restriction enzyme Sau3A I and ligated into Bgl II-digested pIJ702 for the transformation of Streptomyces lividans 1326JI. The primary selection of transformants was made by the observation
of
the
colonies of thiostrepton-resistant clone. From the selected transformants, positive clones of α-1,3 glucanase gene were detected as the clear zones on a modified Czapeck-dox agar medium containing the insoluble glucan as a sole carbon source.
This
transformant possessed a single plasmid, designated pYH1, which contained the vector DNA and a 2.4 Kb Sau3A I insertion fragment was encoded α-1,3 glucanase and was named Streptomyces lividans T1027. The enzyme was partially purified by 30-70%
(NH4)2SO4 precipitation. The cloned enzyme activity was higher and more stable than the enzyme from Streptomyces sp. SW403.
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