He experiments: QZ HC LY HX. Analyzed the data: QZ HC LY HX. Contributed reagents/materials/analysis

He experiments: QZ HC LY HX. Analyzed the data: QZ HC LY HX. Contributed reagents/materials/analysis tools: LY QZ. Wrote the manuscript: QZ.
NIH Public AccessAuthor ManuscriptBiochemistry. Author manuscript; Dihydroorotate Dehydrogenase Inhibitor Storage & Stability accessible in PMC 2014 October 28.Published in final edited kind as: Biochemistry. 2013 April 30; 52(17): 2905?913. doi:10.1021/bi4003343.NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptThe orphan protein bis–glutamylcystine reductase joins the pyridine nucleotide-disulfide reductase familyJuhan Kim1,two and Shelley D. Copley1,two,1Departmentof Molecular, Cellular and Developmental Biology, University of Colorado, Boulder, Boulder, Colorado 80309, United States2CooperativeInstitute for Study in Environmental Sciences, University of Colorado Boulder, Boulder, Colorado 80309, United StatesAbstractFacile DNA sequencing became feasible decades just after a lot of enzymes had been purified and characterized. Consequently, you will discover nevertheless “orphan” enyzmes whose activity is identified however the genes that encode them haven’t been identified. Identification in the genes encoding orphan enzymes is very important because it enables right annotation of genes of unknown function or with mis-assigned function. Bis–glutamylcystine reductase (GCR) is definitely an orphan protein that was purified in 1988. This enzyme catalyzes the reduction of bis–glutamylcystine. Glutamylcysteine (-Glu-Cys) could be the big low molecular weight thiol in halobacteria. We purified GCR from Halobacterium sp. NRC-1 and identified the sequence of 23 tryptic peptides by NanoLC electrospray ionization tandem mass spectrometry. These peptides cover 62 on the protein predicted to be encoded by a gene in Halobacterium sp. NRC-1 which is annotated as mercuric reductase. GCR and mercuric reductase activities were assayed applying enzyme that was expressed in E. coli and re-folded from inclusion bodies. The enzyme had robust GCR activity, but no mercuric reductase activity. The genomes of most, but not all, halobacteria for which entire genome sequences are readily available have close homologs of GCR, suggesting that there is far more to become discovered about the low molecular weight thiols utilised in halobacteria. Enormous genome sequencing efforts in current years have contributed millions of sequences to genomic databases. Functions for the vast majority of these sequences have already been predicted computationally based upon sequence similarities to other proteins along with a wide variety of other genomic clues including genome context and phylogenetic profiling.1? Computational annotations are often correct in the superfamily level. However, predictions of certain functions are generally incorrect. As a result of mis-annotation and subsequent transfer of SphK2 Species erroneous annotations, the database is littered with incorrect assignments of function.four On the other side of your picture, there are a number of “orphan” proteins for which functions are known but for which the corresponding genes haven’t been identified.five? Bis–To whom correspondence needs to be addressed: Shelley D. Copley, Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, Colorado 80309, USA, Tel: (303) 492-6328, Fax: (303) 492-1149, [email protected]. Supplemental Materials may well be accessed cost-free of charge on-line at pubs.acs.org.Kim and CopleyPageglutamylcystine reductase (GCR) is one of these orphan proteins. GCR from Halobacterium halobium was purified and characterized by Sundquist and Fahey in 1988.9 The enzyme.