Albert G. Moat

The relation between penicillin structure and penicillinase activity

Abstract Staphylococcal penicillinase was isolated and purified and its activity and properties were compared with Bacillus penicillinase. Relatively little change was observed in the Michaelis constants and maximum velocities for either enzyme when the phenyl, benzyl, and aliphatic penicillin series were employed as substrates. Compounds with a positively charged nitrogen in the side-chain (e.g., α-aminobenzylpenicillin, p -aminobenzylpenicillin, and 6-aminopenicillanic acid) produced evidence of a lowered affinity for the enzyme. The marked shift in the optimal pH for activity of the staphylococcal enzyme with 6-aminopenicillanic acid provided additional evidence that a decrease in binding results from the presence of a positive charge in the side-chain. With the Staphylococcus enzyme, the Michaelis constants increased with increasing acid strength of the parent side-chain acids when a series of phenyl penicillins was employed, providing evidence for the presence of an electrophilic or positively charged group at the binding site of penicillin to the enzyme. The participation of histidine in the active site of staphylococcal penicillinase was indicated by the pH-activity curve for benzylpenicillin. Methicillin showed a marked decrease in binding to the enzyme with a relatively small change in maximal velocity (about 4000-fold vs. 25-fold). An explanation for the behavior of the enzyme with methicillin is presented.

Factors influencing the synthesis of penicillinase by Micrococcus pyogenes

Abstract Some of the factors influencing synthesis of penicillinase by M. pyogenes are described. Adaptive formation of the enzyme by this organism was not observed under conditions permitting adaptive formation by Bacillus cereus . Penicillinase activity of cells harvested from various media varies considerably suggesting the possibility that certain agents may suppress enzyme formation.

A comparison of nutritional and genetic blocks in the synthesis of purines by yeasts, molds and bacteria☆

Abstract 5-Aminoimidazole riboside and inosine are accumulated by biotin-deficient yeast. The addition of aspartic acid prevents accumulation of these intermediates in the incubation medium. Purine-requiring yeast, neurospora, and vibrio strains accumulate 5-aminoimidazole riboside but not inosine. Aspartic acid fails to prevent 5-aminoimidazole accumulation, an indication of a genetic block in the ability of these organisms to metabolize this compound further. The relationship of biotin to purine biosynthesis by yeast is discussed in the light of the data presented.

Biosynthesis of NAD in Haemophilus haemoglobinophilus

Abstract Haemophilus haemoglobinophilus, a species of Haemophilus which is not dependent upon NAD for growth, was investigated with regard to the pathway of biosynthesis synthesis of this cofactor. Cell-free extracts synthesized NAD directly from nicotinamide without prior deamidation via the following reactions: Nicotinamide+5-phosphoribosyl 1-prrophosphate+ATP → NMN NMN+ATP → NAD Of the enzymes known to be operative in the pathway of NAD biosynthesis from nicotinic acid (nicotinate phosphoribosyltransferase, nicotinate adeninedinucleotide pyrophosphorylase and NAD synthetase) only nicotinate phosphoribosyltransferase activity was demonstrable. Cell free extracts of H. haemoglobinophilus did not degrade NAD and nicotinamidase activity was absent. Quinolic acid was not converted to NAD. The results obtained provided evidence that this organism cannot recycle NAD or synthetize it via the de novo pathway. Direct synthesis from nicotinamide via NMN appears to be the sole pathway of NAD formation in this organism.

Convenient method for enzymic synthesis of 14C-nicotinamide riboside

Abstract A method for the preparation of 14C-nicotinamide riboside from 14C-NAD using a crude enzyme preparation from Proteus vulgaris OX-19 has been described. By heating the preparation in boiling water for 2 min, the enzymes that degrade nicotinamide riboside and nicotinamide are inactivated, providing a system that yields 14C-nicotinamide riboside at 75–80% of the original NAD and with a radiochemical purity greater than 99%.

Effect of Cephalosporin C and Various Penicillin Derivatives on Staphylococcal Penicillinase and Penicillinase-Producing Staphylococcia.∗

Summary and conclusions 1. Cephalosporin C is equally inhibitory to penicillinresistant and penicillin-sensitive staphylococci. However, against sensitive strains it is not nearly so effective an antibiotic as penicillin G. 2. Cephalosporin C and penicillin G apparently do not act synergistically in inhibiting the growth of penicillin-resistant staphylococci. 3. Cephalosporin C, dimethoxyphenylpenicillin and 6-aminopenicillanic acid are hydrolyzed slowly, if at all, by staphylococcal penicillinase. 4. Cephalosporin C is an active inducer of penicillinase synthesis in staphylococci, being comparable to dimethoxyphenylpenicillin in this respect but superior to penicillin G. 5. Cephalosporin C, at relatively high concentrations, does not significantly inhibit staphylococcal penicillinase.

The effect of nutritional factors on the synthesis of staphylococcal penicillinase

Abstract A variety of nutritional factors have been found to exert a marked influence upon the synthesis of penicillinase by staphylococci. Of the amino acids, glutamic acid and tyrosine exerted the greatest stimulatory effect while isoleucine, serine, methionine, and threonine had the greatest inhibitory influence. Of the carbohydrates, maltose raised penicillinase levels threefold over that of glucose. Purines were found to inhibit penicillinase synthesis while pyrimidines did not.