Separation, purification and properties of beta-lactamase I and beta-lactamase II from Bacillus cereus 569/H/9.
1. A procedure was devised which is suitable for the isolation of beta-lactamase I and beta-lactamase II from Bacillus cereus 569/H/9 on a large scale. After adsorption on to Celite both enzymes were eluted in good yield and separated by chromatography on Sephadex CM-50. 2. beta-Lactamase I was separated into three main components by isoelectric focusing and into two components by chromatography. 3. The Zn(2+)-requiring beta-lactamase II obtained by this procedure had a lower molecular weight (22000) than beta-lactamase I (28000) and also differed from the latter in containing one cysteine residue. 4. The beta-lactamase II contained no carbohydrate, but showed the thermostability of the enzyme isolated earlier as a protein-carbohydrate complex. 5. Amino acid analyses and tryptic-digest ;maps' indicate that some degree of homology between beta-lactamase I and beta-lactamase II is possible, but that beta-lactamase I is not composed of the entire sequence of beta-lactamase II together with an additional peptide fragment. 6. A 6-methylpenicillin and a 7-methylcephalosporin showed much lower affinities for both enzymes than did penicillins and cephalosporins themselves.
D000596:Amino Acids / Q000032:analysis; D001409:Bacillus cereus / Q000201:enzymology*; D001431:Bacteriological Techniques; D002241:Carbohydrates / Q000032:analysis; D002511:Cephalosporins; D003470:Culture Media; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D066298:In Vitro Techniques; D008970:Molecular Weight; D010400:Penicillin G; D010405:Penicillinase / Q000032:analysis / Q000302:isolation & purification*; D013696:Temperature; D014462:Ultrafiltration
D000596:Amino Acids; D002241:Carbohydrates; D002511:Cephalosporins; D003470:Culture Media; D010405:Penicillinase; D010400:Penicillin G