Definitive evidence for the discrete biological role of Prolyl. Oligopeptidase (PO) remains unknown, though various physiologi- cal functions for the enzyme have ...
Biochemical Society Transactions (200 I ) Volume 29, Part 5
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Amino acid sequencing and possible cloning of a new proline specific peptidase from bovine serum. Patrick I. Collins and Brendan F. O’Connor School of Biotechnology, Dublin City University, Dublin 9, Eire Definitive evidence for the discrete biological role of Prolyl Oligopeptidase (PO) remains unknown, though various physiological functions for the enzyme have been implicated. Many peptide hormones and neuropeptides are substrates of PO in vitro. Therefore, PO is assumed to be involved in their maturation and degradation in vivo. During our study on bovine serum we have discovered a second PO-like enzyme activity. This enzyme activity was shown to be completely insensitive towards the specific PO inhibitor Z-Pro-prolinal and thus was designated Z-Pro-prolinal insensitive peptidase (ZIP). ZIP activity represents 50% of the apparent PO levels in bovine serum. Using hydrophobic interaction chromatography, we were able to separate the two activities and purify ZIP further through hydroxylapatite and gel filtration chromatography. The N-terminal amino acid sequence of ZIP is presently been sought which will clarify the relation between PO and this second PO-like enzyme activity. Substrate specificity via HPLC and mass spectrophotometry analysis involving proline containing bioactive peptides will further the distinction between these two enzyme activities.The authors acknowledge generous financial support from the Health Research Board and DCU, Eire.
62 Glutamate dehydrogenase from the halophilic organism Halobacterium halobium B.M.Hayden, M.J.Bonete” and P.C. Engel
Department of Biochemistry and Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Ireland. “Department of Biochemistry, University of Alicante, Alicante, Spain. The proteins of halophilic archaea are highly adapted and magnificently engineered to function in high salt. Because of this extreme and unfriendly environment, halophilic (literally salt-loving) proteins and their encoding genes represent a valuable resource for reconstructing processes of natural selection and adaptive evolution. It has previously been reported that the halophilic organism Halobacterium halobium contains both an NAD+ and an NADPf specific glutamate dehydrogenase. The sequence for one of these enzymes has also been reported. In order to gain a deeper understanding of the structural features of the G D H enzymes that are responsible for its halophilic adaptation, we are undertaking the task of isolating the sequenced gene and expressing it in Escherichia coli. We are also attempting to sequence the second G D H gene. Results of the molecular approach are presented.
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Regulation of the serotonin transporter by Syntaxin 1.4 A.M. Killian. F. Magnani, C. Tate, C. Williams, J. Haase. Biochemistry Department, Trinity College Dublin 2, Ireland.
Antidepressant- and cocaine-sensitive serotonin transporter (SERT) functions in the clearance of extracellular serotonin (5-HT) after the neurotransmitter is released. Besides the known protein kinase C (PKC)- mediated regulation of SERT, we explored the regulation of the transporter activity by syntaxin 1A (Syn 1A). Syn 1A as constituent of a SNARE complex is implicated in the docking of synaptic vesicles with the plasma membrane during exocytosis. Futhermore, it appears to interact with and regulate a number of ion channels as well as the SERT-related GABA and glycine transporters. Evidence of interaction between SERT and Syn 1A has been derived from different approaches: (i) the co-expression of Syn 1A and SERT in HEK 293 cells results in a decrease of 5-HT uptake; (ii) confocal microscopy has demonstrated co-localization between SERT and Syn 1A; (iii) Syn 1A was shown to interact with the Nterminal domain of SERT using a GST-pulldown assay. These results suggest that Syn 1A plays a role in the regulation of SERT function.
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Significant raise of IgM and IgG anti-(xanthine oxidoreductase) antibodies in synovial fluid of rheumatoid arthritis patients Lekhmici ARRAR, Nadjet HANACHI, Abderrahmene BAGHIANI and Mustapha BENBOUBETRA
Laboratory of Applied Biochemistry, Faculty Ferhat Abbas of Setif; ALGERIA
of
Sciences, University
Xanthine oxidoreductase (XOR) is a widely distributed molybdoflavo-enzyme that became, over the last two decades, a subject of great interest by its ability to generate reactive oxygen species (ROS) known to be involved in a number of human pathologies. Circulating antibodies to this enzyme have been, initially, reported some twenty five years ago and confirmed in a number of papers. We have already reported the presence of such antibodies in healthy normal humans. The origin of human anti-XOR antibodies is still unclear despite the fact that our previous results strongly point out to the endogenous XOR as the immunogen rather than the ingested cow’s milk enzyme. We report here, an ELISA detection of elevated anti-XOR antibodies titres in synovial fluid from patients with rheumatoid arthritis, chronic arthritis and from patients with other joint inflammations. As far as we are aware of, the presence of such high levels of anti-XOR antibodies in synovial fluid has not been reported before. The interference of rheumatoid factor in the assay has been thoroughly investigated. A high proportion of synovial anti-XOR antibodies forms immune complexes. These antibodies could well play a protective role by complexing the ROS generating enzyme, XOR, but the possibility of immune complexe diseases, by the build up of immune complexes in joints, can not be ruled out.
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Biochemical Society
