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Advances in Applied Biotechnology Part 10

Tham khảo tài liệu 'advances in applied biotechnology part 10', khoa học tự nhiên, công nghệ sinh học phục vụ nhu cầu học tập, nghiên cứu và làm việc hiệu quả | Approaches for Improving Protein Production in Multiple Protease-Deficient Bacillus Subtilis Host Strains 169 degradation of a-amylase-A522-PreS2 by the inactivation of IspA in the KA8AX strain. However the productivity of a-amylase-A522-PreS2 in the ten-protease deficient mutant was almost same as that in the KA8AX strain. A search in the GenoList database for B. subtilis 168 genome http genodb.pasteur.fr cgi-bin WebObjects GenoList of proteases and peptidases revealed the presence of 31 known and 11 putative proteases and 38 known and 12 putative peptidases respectively. Section 2.2 describes the investigation of membrane-bound proteases involved in protein degradation. Fig. 5. Western blot analysis of the a-amylase-A522-PreS2 hybrid protein in the extracellular fractions of Dpr7 Dpr8 and KA8AX. A Western blot analysis was carried out to detect a-amylase-A522-PreS2 with the anti-PreS2 antibody. Culture supernatants from Dpr7 lanes 1-3 Dpr8 lanes 4-6 and KA8AX lanes 7-9 were collected after 25 50 h and 75 h of cultivation and subjected to Tricine-SDS-PAGE and Western blotting as described in the Materials and Methods. Proteins from the culture supernatants equivalent to 1 pl were applied to each lane. The arrowhead indicates the position of a-amylase-A522-PreS2. The times of harvest of supernatants are shown at the top. B The relative a-amylase-A522-PreS2 protein amounts were compared on the basis of band intensities on Western blots the amount of a-amylase-A522-PreS2 at 50 h in the Dpr8 strain was set to 100 . The presented results are the average of three individual experiments. Error bars correspond to the standard errors of the means SEM . Lane numbers in panel A correspond to those in panel B. 170 Advances in Applied Biotechnology Fig. 6. The a-amylase-A522-PreS2 hybrid protein was degraded by AprX. A Zymography of supernatants from the KA8AX pDG-AprX strain. Lane 1 without IPTG lane 2 with IPTG. B Western blot analysis of degradation of a-amylase-A522-PreS2