Cloned penicillin G acylase (PGA) from Escherichia coli ATCC 11105 was mutagenized in vivo using N-methyl-N'-nitro-N-nitrosoguanidine. Mutants of PGA were selected by their ability to allow growth of the host strain E. coli M8820 with the new substrates phenylacetyl-beta-alanyl-L-proline (PhAc-beta Ala-Pro) phthalyl-L-leucine (Pht-Leu) or phthalylglycyl-L-proline (Pht-Gly-Pro) as sole source of proline and leucine respectively. PGA mutants were purified and immobilized onto spherical methacrylate (G-gel). The immobilized form of mutant PGA selected with (PhAc-beta Ala-Pro) hydrolyzed 95% of 9 mmol penicillin G 30% faster than wild-type PGA using the same specific activities. The specific activity of the soluble enzyme was 2.7-fold, and inhibition by phenylacetic acid was halved. Immobilized PGA mutant selected with Pht-Gly-Pro hydrolyzed penicillin G 20% faster than wild-type PGA. The Km of the soluble enzyme was increased 1.7-fold. Furthermore, the latter two mutants were also 3.6-fold more stable at 45 degrees C than wild-type PGA. The specific activity of the mutant selected with Pht-Leu was 6.3-fold lower, and inhibition by phenylacetic acid was increased 13-fold.

Enzyme Stability↗

PubMedDate not supplied

[BCG-vaccination in neonates as a cause of osteomyelitis--an argument against BCG-vaccination?].

BCG-osteomyelitis is a rare complication of BCG-vaccination. By means of a case report clinical and immunological findings are discussed. Contrary to the undramatic clinical symptoms the x-ray showed pronounced osteolytic lesions in the sternum. The bacteriological examination demonstrated mycobacteria of BCG-type. Immunological investigations presented normal humoral and cellular immunity. The therapy (curettage, antituberculous drugs) was effective. In spite of this complication the BCG-vaccination is furthermore recommended.

BCG Vaccine↗