TAILIEUCHUNG - Báo cáo khóa học: Effects of Escherichia coli ribosomal protein S12 mutations on cell-free protein synthesis

We examined the effects ofEscherichia coliribosomal pro-tein S12 mutations on the efficiency of cell-free protein syn-thesis. By screening 150 spontaneous streptomycin-resistant isolates from E. coliBL21, we successfully obtained seven mutants of the S12 protein, including two streptomycin-dependent mutants. The mutations occurred at Lys42, Lys87, Pro90 and Gly91 of the 30S ribosomal protein S12. We prepared S30 extracts frommutant cells harvested in the mid-log phase. | Eur. J. Biochem. 271 1127-1134 2004 FEBS 2004 doi Effects of Escherichia coli ribosomal protein S12 mutations on cell-free protein synthesis Namthip Chumpolkulwong1 Chie Hori-Takemoto1 Takeshi Hosaka2 Takashi Inaoka2 Takanori Kigawa1 Mikako Shirouzu1 3 Kozo Ochi2 and Shigeyuki Yokoyama1 3 4 1RIKEN Genomic Sciences Center Tsurumi Yokohama Japan 2National Food Research Institute Tsukuba Ibaraki Japan 3RIKEN Harima Institute at SPring-8 Mikazuki-cho Sayo Hyogo Japan 4Department of Biophysics and Biochemistry Graduate School of Science The University of Tokyo Bunkyo-ku Tokyo Japan We examined the effects of Escherichia coli ribosomal protein S12 mutations on the efficiency of cell-free protein synthesis. By screening 150 spontaneous streptomycin-resistant isolates from E. coli BL21 we successfully obtained seven mutants of the S12 protein including two streptomycindependent mutants. The mutations occurred at Lys42 Lys87 Pro90 and Gly91 of the 30S ribosomal protein S12. We prepared S30 extracts from mutant cells harvested in the mid-log phase. Their protein synthesis activities were compared by measuring the yields of the active chloramphenicol acetyltransferase. Higher protein production than the wild-type was observed with the mutant that replaced Lys42 with Thr K42T . The K42R K42N and K42I strains showed lower activities while the other mutant strains with Lys87 Pro90 and Pro91 did not show any significant difference from the wild-type. We also assessed the frequency of Leu misincorporation in poly U -dependent poly Phe synthesis. In this assay system almost all mutants showed higher accuracy and lower activity than the wild-type. However K42T offered higher activity in addition to high accuracy. Furthermore when 14 mouse cDNA sequences were used as test templates the protein yields of nine templates in the K42T system were times higher than that of the wild-type. Keywords ribosomal protein S12 streptomycin point .

• Báo cáo khoa học
• Report medicine
• traditional medicine
• scientific reports
• intermediate cells
• cell proliferation
• medical analysis
• medical knowledge
• medical research
• analysis of cytoplasmic
• oxidation
• Crystal structure
• bacteria
• hemoglobin
• medicine
• breast cancer
• chemical reaction
• gastric cancer
• hormone medicine
• tumor cells
• biochemical studies
• environmental medicine