Disease relevance of Nitrocefin

• Immunofluorescence microscopy, the accessibility of the fusion protein to externally added proteases, and the rates of hydrolysis of nitrocefin and penicillin G by whole cells demonstrated that a substantial fraction (20-30%) of the beta-lactamase domain of the fusion protein was exposed on the external surface of E. coli [1].
• The catalytic mechanism of metallo-beta-lactamase from Bacteroides fragilis, a dinuclear Zn(II)-containing enzyme responsible for multiple antibiotic resistance, has been investigated by using nitrocefin as a substrate [2].
• Hydrolysis of the chromogenic beta-lactam nitrocefin by periplasmic beta-lactamase in intact Pseudomonas aeruginosa cells was used to assess the influence of various compounds on the permeability of the P. aeruginosa outer membrane [3].
• The reaction of nitrocefin with metallo-beta-lactamase L1 from Stenotrophomonas maltophilia was studied using rapid-scan and stopped-flow ultraviolet-visible (UV-vis) studies in an effort to discern the kinetic mechanism used by L1 to hydrolyze penicillins and cephalosporins [4].
• The prevalence of beta-lactamase-producing bacteria in subgingival plaque from patients with refractory periodontitis in Norway was assessed by the chromogenic nitrocefin method. beta-Lactamase activity was detected in 68% of the patients [5].

Psychiatry related information on Nitrocefin

• S1 reaction times were approximately 50% faster than nitrocefin for beta-lactamase-positive enterococci and S. aureus [6].

High impact information on Nitrocefin

• In cells grown at 24 degrees C the localization of beta-lactamase on the cell surface was almost quantitative (greater than 80% of the enzymatically active protein was exposed to the extracellular fluid) as determined by nitrocefin and penicillin G hydrolysis and trypsin accessibility [1].
• In the case of D120(81)E, the nitrocefin hydrolysis product, which shows a maximum absorption at 460 nm, was bound to D120(81)E in the protonated form [7].
• Additionally, the 2.4-angstroms structure of the acyl-enzyme complex of R39 with nitrocefin reveals the absence of active site conformational change upon binding the beta-lactams [8].
• We suggest, therefore, that the mechanism of hydrolysis of nitrocefin by binuclear metallo-beta-lactamases may be atypical and that cleavage of the beta-lactam amide bond is the rate-determining step for breakdown of the majority of beta-lactam substrates by the L1 enzyme [9].
• The 200 ps QM/MM simulation of the CcrA enzyme in complex with nitrocefin shows that the substrate beta-lactam moiety is directed toward the active site dizinc center through the interactions of aminocarbonyl and carboxylate groups with the two active site zinc ions and the two conserved residues, Lys167 and Asn176 [10].

Chemical compound and disease context of Nitrocefin

• It has been presumed that there are just two beta-lactamases in the motile Aeromonas species, a carbapenemase and a cephalosporinase, based on the premise that all beta-lactamases can be detected by hydrolysis of the chromogenic cephalosporin, nitrocefin [11].
• Plasmid-encoded beta-lactamases were rare in Klebsiella spp., being found in six (7.5%) isolates of K. pneumoniae and in none of the K. oxytoca. beta-Lactamase activities were relatively low (< 100 nmoles nitrocefin hydrolysed per minute per mg of protein) and ampicillin MICs were < or = 128 mg/L for most isolates of both species [12].
• A substrate, pyridine-2-azo-p-dimethylaniline cephalosporin (PADAC; Diagnostic Pasteur, Marnes-La-Coquette, France), for detection of penicillinase-producing Neisseria gonorrhoeae (PPNG) on isolated colonies grown on agar was compared with the nitrocefin reference test (Cefinase; Biomerieux, Marcy l'Etoile, France) [13].
• Extended X-ray absorption fine structure studies of the metallo-beta-lactamase L1 from Stenotrophomonas maltophilia containing 1 and 2 equiv of Zn(II) and containing 2 equiv of Zn(II) plus hydrolyzed nitrocefin are presented [14].
• Twenty micromolar tazobactam inhibited total beta-lactamase activity (as measured by nitrocefin hydrolysis rates) by greater than 75% against all isolates tested: in 11 of 13 E. coli isolates, total beta-lactamase activity was inhibited by 90% [15].

Biological context of Nitrocefin

• Using nitrocefin, a chromogenic beta-lactam, we confirmed the correlation between PBP 2a' precipitation and its beta-lactam-dependent enzymatic acylation by monitoring the absorbance associated with the precipitate [16].
• IC50 values (microM) with nitrocefin, piperacillin and penicillin G substrates (concentrations 20, 100 and 20 microM, respectively) were > 1000, 7, 3.5 (clavulanate); > 1000, 300, 59 (sulbactam), > 1000, 29, 7.7 (tazobactam); 0.0004, 0.001, 0.0018 (BRL 42715) [17].
• The kinetics of the La(3+)-catalyzed methanolysis of N-phenyl-beta-lactam (2) and N-p-nitrophenyl-beta-lactam (3) as well as that of nitrocefin (1) were studied at 25 degrees C under buffered conditions [18].
• Cefoxitin susceptibility was performed using an agar dilution method, beta-lactamase production by using a nitrocefin method, and plasmid extraction by using a commercial kit [19].
• Three different beta-lactamase substrate profiles were identified in 95 isolates of coagulase-negative staphylococci (CNS), of 57 different phenotypes, from 16 orthopaedic inpatients and staff members in one ward, by applying a bacterial whole-cell assay based on the hydrolysis of cefazolin, cephaloridine and nitrocefin [20].

Anatomical context of Nitrocefin

• A colorimetric method, based on the conversion of nitrocefin, was used for quantitation of the sputum beta-lactamase activity. beta-Lactamases in sputum were characterized by isoelectric focusing and inhibition profile and were compared with the beta-lactamases extracted from Pseudomonas aeruginosa isolated from the paired sputum samples [21].
• Neither of these strains showed evidence of decreased permeability to nitrocefin as judged by a method which takes into account the substrate concentration in the periplasmic space [22].
• The chromogenic cephalosporin nitrocefin was used to detect the presence of beta-lactamase in cerebrospinal fluid of patients suffering from ampicillin-resistant Haemophilus influenzae meningitis [23].
• Different mesophilic methanogenic and extremely halophilic Archaea containing methanochondroitin, pseudomurein, or S-layer protein or glycoprotein cell walls, were tested for beta-lactamase activity with the chromogenic beta-lactam nitrocefin as substrate [24].
• METHODS: beta-Lactamase activity in culture supernatants and in cytoplasmic membrane fractions was estimated by bioassay and by SDS-PAGE combined with nitrocefin assay [25].

Associations of Nitrocefin with other chemical compounds

• Partial screening of this bifunctional sublibrary resulted in the identification of two enzymes in which the presence of maltose modulated the rate of nitrocefin hydrolysis [26].
• Preliminary experiments led to the selection of suitable systems for the study of beta-lactamase II catalysis at low temperatures, namely, cobalt(II) beta-lactamase II hydrolysis of benzylpenicillin in 60% (v/v) ethylene glycol and zinc beta-lactamase II hydrolysis of the chromophoric cephalosporin nitrocefin in 60% (v/v) methanol [27].
• The hydrolysis of nitrocefin by zinc beta-lactamase II has been studied concurrently in 60% (v/v) methanol [27].
• It exhibits no detectable activity toward benzylpenicillin, and 10(5)-fold reduction of kcat for nitrocefin hydrolysis compared with the wild-type enzyme [28].
• Furthermore, the inhibition of the beta-lactamase hydrolysis of nitrocefin by cefoxitin or moxalactam was prevented by a 12-h preincubation of the beta-lactam with the B. bivius extracts but not with the B. fragilis extracts [29].

Gene context of Nitrocefin

• Testing beta-lactamase production with nitrocefin was more predictive for the presence of the blaZ gene than the agar dilution method and the results of the former agreed highly with the presence of the blaZ gene in the isolates [30].
• Compounds 3a-c, 7a-c were able to inhibit either TEM-1 (a Class A enzyme, from Escherichia coli) or P-99 (a Class C enzyme, from E. cloacae), or both enzymes, when tested in competition experiments using nitrocefin as the reporter substrate [31].
• Using a nitrocefin competition assay, I determined the relative substrate affinity index (RSAI) values of nine clinically significant beta-lactamase enzymes against a range of beta-lactams [32].
• This was particularly true for the bile-resistant or Bacteroides fragilis group. beta-Lactamase production was detected in 82% of the bacteroides with the nitrocefin test [33].
• Familial mutations and zinc stoichiometry determine the rate-limiting step of nitrocefin hydrolysis by metallo-beta-lactamase from Bacteroides fragilis [34].

Analytical, diagnostic and therapeutic context of Nitrocefin

• The role of the mobile loop comprising residues 60-66 in metallo-beta-lactamases has been studied by site-directed mutagenesis, determination of kinetic parameters for six substrates and two inhibitors, pre-steady-state characterization of the interaction with chromogenic nitrocefin, and molecular modeling [35].
• Nitrocefin tests were performed to detect the beta-lactamase enzyme; isoelectric focusing electrophoresis, PCR, and PCR product sequencing were designed to further characterize the contained ESBLs [36].
• PADAC exhibited little antimicrobial activity against gram-negative bacteria, but did have good activity (minimum inhibitory concentration, 0.12 to 0.5 microgram/ml) against Staphylococcus aureus, a quality comparable to nitrocefin [37].
• Beta-lactamases were examined with a conventional nitrocefin colony testing procedure, activity and induction assays, and SDS-PAGE [38].
• CONCLUSION--Detection of beta lactamase activity in sputum using nitrocefin, after treatment with blue sepharose beads, is a rapid, reproducible test with high specificity and sensitivity [39].

References