Doug D Buechter

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Epoxycreatine (N-(2,3-epoxypropyl)-N-amidinoglycine) is an affinity label of creatine kinase that irreversibly and completely inactivates the enzyme (Marletta, M. A., and Kenyon, G. L. (1979) J. Biol. Chem. 254, 1879-1886). To identify active site residues of rabbit muscle creatine kinase, the site of modification of it by epoxycreatine has been determined.(More)
The genes that encode the five known enzymes of the mandelate pathway of Pseudomonas putida (ATCC 12633), mandelate racemase (mdlA), (S)-mandelate dehydrogenase (mdlB), benzoylformate decarboxylase (mdlC), NAD(+)-dependent benzaldehyde dehydrogenase (mdlD), and NADP(+)-dependent benzaldehyde dehydrogenase (mdlE), have been cloned. The genes for(More)
Expression of creatine kinase (CK) from a Torpedo californica electric organ cDNA in Escherichia coli results in an insoluble protein product with no detectable CK activity. Although this is a stable aggregate that can be isolated in an enriched form by centrifugation, initial attempts to generate enzyme activity by denaturing and refolding yielded only(More)
The genetic code is based on the aminoacylation of tRNA with amino acids catalyzed by the aminoacyl-tRNA synthetases. The synthetases are constructed from discrete domains and all synthetases possess a core catalytic domain that catalyzes amino acid activation, binds the acceptor stem of tRNA, and transfers the amino acid to tRNA. Fused to the core domain(More)
The catabolic cytokine interleukin-1 (IL-1) and endotoxin lipopolysaccharide (LPS) are well-known inflammatory mediators involved in degenerative disc disease, and inhibitors of IL-1 and LPS may potentially be used to slow or prevent disc degeneration in vivo. Here, we elucidate the striking anti-catabolic and anti-inflammatory effects of bovine(More)
OBJECTIVE Protein kinase Cδ (PKCδ) activation has been shown to be a principal rate-limiting step in matrix-degrading enzyme production in human articular chondrocytes. The aim of this study was to assess the role of the PKC pathways, specifically PKCδ, in intervertebral disc tissue homeostasis. METHODS Using in vitro, ex vivo, and in vivo techniques, we(More)
A core of eight beta-strands and three alpha-helices was recently predicted for the active site domain of Escherichia coli alanyl-tRNA synthetase, an enzyme of unknown structure [Ribas de Pouplana, L1., Buechter, D. D., Davis, M. W., & Schimmel, P. (1993) Protein Sci. 2, 2259-2262; Shi, J.-P., Musier-Forsyth, K., & Schimmel, P. (1994) Biochemistry 26,(More)
The ten class II aminoacyl-tRNA synthetases are large homo- and hetero-oligomeric proteins that share three conserved sequence motifs. Within this class, Escherichia coli alanyl-tRNA synthetase is the only homotetramer and is comprised of subunits of 875 amino acids. The enzyme aminoacylates sequence-specific RNA oligonucleotides that recreate as few as(More)
The class-defining active site domain of the 10 class II tRNA synthetases is well conserved and, based on the crystal structure of aspartyl-tRNA synthetase, approaches the end of the tRNA acceptor stem from the major groove side of the helix. Paradoxically, for the class II alanyl-tRNA synthetase (AlaRS), aminoacylation is dependent on minor groove(More)