Biomaterial Database

Purification of beta-lactamase from Streptomyces cellulosae by affinity chromatography on Blue Sepharose. 1979

H Ogawara, and S Horikawa

A beta-lactamase from culture supernatant of Streptomyces cellulosae was purified about 1,450-fold to apparent homogeneity in polyacrylamide gel electrophoresis and isoelectric focusing on polyacrylamide gel sheet. The methods used were ammonium sulfate precipitation, CM-52 cellulose ion-exchange chromatography and affinity chromatography on Blue Sepharose CL-6B. The molecular weight was determined to be approximately 27,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This value was in good agreement with the previous value determined by gel filtration on Sephadex G-75. The isoelectric point was pH 9.5. The enzyme behaved primarily as penicillinase and apparent Km value for benzylpenicillin was 500 microM. The beta-lactamase of S. cellulosae interacted strongly with blue dextran and NADP+-agarose but not with Sepharose. In addition, the presence of NADP+ but not NAD+ and ATP diminished sharply the intrinsic fluorescence intensity of the enzyme and the apparent association constant was calculated to be 1.4 x 10(3) M-1. The beta-lactamase decreases its enzymatic activity against benzylpenicillin in the presence of NADP+. From these results, it is suggested that this beta-lactamase has a dinucleotide binding fold.

UI	MeSH Term	Description	Entries
D008722	Methods	A series of steps taken in order to conduct research.	Techniques,Methodological Studies,Methodological Study,Procedures,Studies, Methodological,Study, Methodological,Method,Procedure,Technique
D009711	Nucleotides	The monomeric units from which DNA or RNA polymers are constructed. They consist of a purine or pyrimidine base, a pentose sugar, and a phosphate group. (From King & Stansfield, A Dictionary of Genetics, 4th ed)	Nucleotide
D002627	Chemistry, Physical	The study of CHEMICAL PHENOMENA and processes in terms of the underlying PHYSICAL PHENOMENA and processes.	Physical Chemistry,Chemistries, Physical,Physical Chemistries
D002846	Chromatography, Affinity	A chromatographic technique that utilizes the ability of biological molecules, often ANTIBODIES, to bind to certain ligands specifically and reversibly. It is used in protein biochemistry. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)	Chromatography, Bioaffinity,Immunochromatography,Affinity Chromatography,Bioaffinity Chromatography
D000596	Amino Acids	Organic compounds that generally contain an amino (-NH2) and a carboxyl (-COOH) group. Twenty alpha-amino acids are the subunits which are polymerized to form proteins.	Amino Acid,Acid, Amino,Acids, Amino
D001426	Bacterial Proteins	Proteins found in any species of bacterium.	Bacterial Gene Products,Bacterial Gene Proteins,Gene Products, Bacterial,Bacterial Gene Product,Bacterial Gene Protein,Bacterial Protein,Gene Product, Bacterial,Gene Protein, Bacterial,Gene Proteins, Bacterial,Protein, Bacterial,Proteins, Bacterial
D001618	beta-Lactamases	Enzymes found in many bacteria which catalyze the hydrolysis of the amide bond in the beta-lactam ring. Well known antibiotics destroyed by these enzymes are penicillins and cephalosporins.	beta-Lactamase,beta Lactamase,beta Lactamases
D012685	Sepharose		Agarose,Sepharose 4B,Sepharose C1 4B,4B, Sepharose C1,C1 4B, Sepharose
D013302	Streptomyces	A genus of bacteria that form a nonfragmented aerial mycelium. Many species have been identified with some being pathogenic. This genus is responsible for producing a majority of the ANTI-BACTERIAL AGENTS of practical value.
D013379	Substrate Specificity	A characteristic feature of enzyme activity in relation to the kind of substrate on which the enzyme or catalytic molecule reacts.	Specificities, Substrate,Specificity, Substrate,Substrate Specificities