Characterization of the neisserial lipid-modified azurin bearing the H.8 epitope

The pathogenic Neisseria have multiple genes encoding proteins that bind monoclonal antibody (MAb) H.8. We previously reported the cloning and sequencing of a meningococcal gene (laz) encoding an H.8 MAb-binding protein with a consensus lipoprotein processing site, an N-terminal domain containing the epitope for H.8 MAb binding, and a C-terminal domain with extensive similarity to the sequences of azurins from other organisms. In the current study, we showed that the product of the cloned gene could be labelled with palmitic acid, that it was subject to globomycin-sensitive processing, and that it was immunologically cross-reactive with azurin from Pseudomonas aeruginosa. All neisserial species tested, both pathogens and commensals, produced a protein recognized by anti-azurin serum. Southern blots with oligonucleotide probes specific for the azurin domain of the gene showed that it was present in a single copy in the chromosome; it was highly conserved in gonococci and meningococci, and less conserved in commensal Neisseria species.     

A COMPARATIVE INVESTIGATION OF PEROXIDASES FROM GERMINATING PEANUTS (ARACHIS HYPOGAEA): ELECTROPHORESIS

Dormant seed and organs of 0-, 1-, 2-, 5-, 8-, 11-, and 14-day-old plants of Arachis hypogaea L. were homogenized in phosphate buffer and the lipid-free extracts analyzed for benzidine and pyrogallol peroxidases using starch-gel electrophoresis. On a wet weight basis, one weak band of benzidine peroxidase activity was detected in dormant cotyledons and three bands in 1-day cotyledons. In 5-day tissue, activity had increased significantly; at 14 days, the number of bands had decreased but staining intensity was maintained. In the extract from dormant axis, a single cathodic site of benzidine peroxidase activity was observed; however, on day two there was a marked increase in the number of bands and intensity of reaction in epicotyl and hypocotylradicle tissues. By day 14, the number and density of bands had decreased noticeably in the epicotyl and hypocotyl. Extracts from 14-day roots exhibited more sites of reaction and greater intensity of staining of benzidine peroxidase than at five days of growth. Localized areas of activity at Rf -0.44 and -0.52 were present in extracts of all four organs when either benzidine or pyrogallol was used as the hydrogen donor. Although marked similarity existed between banding patterns of organs, qualitative and quantitative ontogenetic differences in peroxidases were apparent.     

ELECTROPHORETIC ANALYSIS OF ENZYMES FROM THREE SPECIES OF CHLAMYDOMONAS

Isoenzyme patterns of acetone-extracted proteins revealed a close similarity between Chlamydomonas eugametos and C. moewusii but a distant relationship between the two and C. reinhardtii. Chlamydomonas eugametos and C. moewusii had identical banding patterns of malate dehydrogenase (MDH) and leucine aminopeptidase (LAP) in starch gels. These two species exhibited the same MDH distribution spectrum in analytical disc polyacrylamide gels but neither species showed definitive LAP or glutamate dehydrogenase (GDH) activity. There were differences in the starch gel alpha esterase (α-EST) patterns of C. eugametos and C. moewusii due to an additional weak band at Rf 0.75 in the latter species and a slight variation in the position of another band at Rf 0.80–0.82. Some variations between the two species also occurred in the α-EST banding in disc gels at Rf 0.70–0.85 and at Rf 0.06–0.14 with C. moewusii exhibiting the greatest number of bands. Chlamydomonas reinhardtii displayed patterns of all four enzymes but the band characteristics were distinctively different from those of C. eugametos and C. moewusii. There appeared to be no obvious isoenzyme difference between mating types of either species. It is concluded that C. eugametos and C. moewusii are not identical species but are closely related in regard to the enzymes assayed. Isoenzyme analysis is considered to be a useful approach to algal systematics.