Serine-Type D-Ala-D-Ala Peptidases and Penicillin-Binding Proteins

in Methods in Enzymology (1994), 244

The Mechanism of Action of DD-Peptidases: The Role of Tyrosine-159 in the Streptomyces R61 DD-Peptidase

in Biochemical Journal (1993), 291((Pt 2)), 537-44

Tyrosine-159 of the Streptomyces R61 penicillin-sensitive DD-peptidase was replaced by serine or phenylalanine. The second mutation yielded a very poorly active protein whose rate of penicillin binding ... [more ▼]

Tyrosine-159 of the Streptomyces R61 penicillin-sensitive DD-peptidase was replaced by serine or phenylalanine. The second mutation yielded a very poorly active protein whose rate of penicillin binding was also drastically decreased, except for the reactions with nitrocefin and methicillin. The consequences of the first mutation were more surprising, since a large proportion of the thiolesterase activity was retained, together with the penicillin-binding capacity. Conversely, the peptidase properties was severely affected. In both cases, a drastic decrease in the transferase activity was observed. The results are compared with those obtained by mutation of the corresponding residue in the class A beta-lactamase of Streptomyces albus G. [less ▲]

Acyltransferase activities of the high-molecular-mass essential penicillin-binding proteins

in Biochemical Journal (1991), 279(Part 2), 601-604

The high-molecular-mass penicillin-binding proteins (HMM-PBPs), present in the cytoplasmic membranes of all eubacteria, are involved in important physiological events such as cell elongation, septation or ... [more ▼]

The high-molecular-mass penicillin-binding proteins (HMM-PBPs), present in the cytoplasmic membranes of all eubacteria, are involved in important physiological events such as cell elongation, septation or shape determination. Up to now it has, however, been very difficult or impossible to study the catalytic properties of the HMM-PBPs in vitro. With simple substrates, we could demonstrate that several of these proteins could catalyse the hydrolysis of some thioesters or the transfer of their acyl moiety on the amino group of a suitable acceptor nucleophile. Many of the acyl-donor substrates were hippuric acid or benzoyl-D-alanine derivatives, and their spectroscopic properties enabled a direct monitoring of the enzymic reaction. In their presence, the binding of radioactive penicillin to the PBPs was also inhibited. [less ▲]