Overproduction and crystallization ofFokI restriction endonuclease
نویسندگان
چکیده
منابع مشابه
Crystallization and preliminary X-ray analysis of restriction endonuclease FokI bound to DNA.
FokI is a type IIs restriction endonuclease which recognizes an asymmetric DNA sequence and cleaves DNA a short distance away from the sequence. The enzyme is bipartite in nature with its DNA recognition and cleavage functions located on distinct domains. We report here cocrystals of the complete FokI enzyme (579 amino acids) bound to a 20-bp DNA fragment containing its recognition sequence. Th...
متن کاملChimeric restriction endonuclease.
Fok I restriction endonuclease recognizes the nonpalindromic pentadeoxyribonucleotide 5'-GGATG-3'.5'-CATCC-3' in duplex DNA and cleaves 9 and 13 nt away from the recognition site. Recently, we reported the presence of two distinct and separable domains within this enzyme: one for the sequence-specific recognition of DNA (the DNA-binding domain) and the other for the endonuclease activity (the c...
متن کاملOverexpression, purification and crystallization of BamHI endonuclease.
The type II restriction endonuclease BamHI has been expressed in E. coli, producing 100-fold more enzyme than the wild type Bacillus amyloliquefaciens H strain. This high yield has facilitated purification to homogeneity of large amounts of the enzyme, along with its crystallization in a form which diffracts to at least 1.9 A in X-ray analysis.
متن کاملOverproduction of the EcoR V endonuclease and methylase.
Strains overproducing the EcoR V endonuclease and methylase have been obtained by inserting each of the two genes in expression vectors containing the lambda PL promoter. The methylase is overproduced to a level reaching 5-10% of the total cellular proteins, which represents a 50-100 fold increase. A 30 fold overproduction of endonuclease was achieved by randomly positioning the EndRV gene down...
متن کاملIsolation and identification of restriction endonuclease BsiSI.
BseBI, an isoschizomer of BstNI (1) has been purified from Bacillus stearothermopilus species. BseBI recognises the sequence 5'...CCWGG...3' (W=A or T) and cleaves between C and W. The enzyme was purified using the following Chromatographic steps: 1. Blue -Sepharose F3GA, 2. Heparin -Sepharose, 3. DEAE-Sepharose. The enzyme was free of contaminating nuclease activity. After 5-fold overdigestion...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
ژورنال
عنوان ژورنال: Nucleic Acids Research
سال: 1989
ISSN: 0305-1048,1362-4962
DOI: 10.1093/nar/17.21.8741