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Oxyanion Hole (oxyanion + hole)
Selected AbstractsFrom Theozymes to Artificial Enzymes: Enzyme-Like Receptors for Michael Additions with Oxyanion Holes and Active Amino GroupsEUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 29 2007Luis Simón Abstract Different artificial enzymes, based on the theozyme concept, have been designed for Michael additions of pyrrolidine to ,,,-unsaturated lactams. These molecules each have skeleton able to mimic a structure called an "oxyanion hole", as is present in many enzymes. Amine groups are also responsible for the catalytic activities of these receptors, since they support the important proton-transport step. The requirement for the amine groups was established from the reaction mechanism and from theoretical calculations. The catalytic activities of the receptors are discussed, taking into account their relative association constants with the substrate: kcat/kuncat values of up to 104 were obtained. The catalytic activities of the receptors are compared with those found in natural enzymes and catalytic antibodies. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Isolation of a low-temperature adapted lipolytic enzyme from uncultivated micro-organismJOURNAL OF APPLIED MICROBIOLOGY, Issue 1 2008C. Roh Abstract Aims:, The aim of the study was to isolate a novel lipolytic enzyme from the activated sludge of uncultured micro-organisms. Methods and Results:, The metagenomic DNA was directly extracted from the activated sludge, and a metagenomic library was constructed by using the pUC vector. The library was screened for lipolytic enzyme activity on 1% tributyrin agar plate. A clone among c. 100 000 recombinant libraries showed the lipolytic activity. The putative lipolytic gene encoding lipo1 from the metagenomic library was subcloned and expressed in Escherichia coli BL21 using the pET expression system. The expressed recombinant enzyme was purified by Ni-nitrilotriacetic acid affinity chromatography and characterized using general substrates of lipolytic property. The gene consisted of 972 bp encoding a polypeptide of 324 amino acids with a molecular mass of 35·6 kDa. Typical residues essential for lipolytic activity such as penta-peptide (GXSXG) and catalytic triad sequences (Ser166, Asp221 and His258) were detected. The deduced amino acid sequence of lipo1 showed low identity with amino acid sequences of esterase/lipase (32%, ZP_01528487) from Pseudomonas mendocina ymp and esterase (31%, AAY45707) from uncultured bacterium. This lipolytic enzyme exhibited the highest activity at pH 7·5 and 10°C. At thermal stability analysis, lipo1 was more unstable at 40°C than 10°C. Conclusions:, An activity based strategy has been an effective method for fishing out a low-temperature adapted lipolytic enzyme from the metagenomic library. This lipo1 enzyme can be considered to belong to the hormone-sensitive lipase family due to the enzyme's oxyanion hole by the sequence HGGG. Significance and Impact of the Study:, Lipo1 is a novel psychrophilic esterase obtained directly from the metagenomic library. Owing its support of significant activity at low temperature, this enzyme is expected to be useful for potential application as a biocatalyst in organic chemistry. [source] Human rhinovirus 3C protease: a cysteine protease showing the trypsin(ogen)-like foldBIOCHEMISTRY AND MOLECULAR BIOLOGY EDUCATION, Issue 4 2001Martino Bolognesi Abstract Viral-encoded proteases cleave precursor polyprotein(s) leading to maturation of infectious virions. Strikingly, human rhinovirus 3C protease shows the trypsin(ogen)-like serine protease fold based on two topologically equivalent six-stranded ,-barrels, but displays residue Cys147 as the active site nucleophile. By contrast, papain, which is representative of most cysteine proteases, does not display the trypsin(ogen)-like fold. Remarkably, in human rhinovirus 3C cysteine protease, the catalytic residues Cys147, His40 and Glu71 are positioned as Ser195, His57 and Asp102, respectively, building up the catalytic triad of serine proteases in the chymotrypsin-trypsin-elastase family. However, as compared to trypsin-like serine proteases and their zymogens, residue His40 and the oxyanion hole of human rhinovirus 3C cysteine protease, both key structural components of the active site, are located closer to the protein core. Human rhinovirus 3C cysteine protease cleaves preferentially Gin , Gly peptide bonds or, less commonly, the Gin , Ser, Gin ,Ala, Glu , Ser or Glu ,Gly pairs. Finally, human rhinovirus 3C cysteine protease and the 3CD cysteine protease-polymerase covalent complex bind the 5° non-coding region of rhinovirus genomic RNA, an essential function for replication of the viral genome. © 2001 IUBMB. Published by Elsevier Science Ltd. All rights reserved. [source] Outer-membrane phospholipase A: known structure, unknown biological functionMOLECULAR MICROBIOLOGY, Issue 4 2000MicroReview Outer-membrane phospholipase A (OMPLA) is one of the few enzymes present in the outer membrane of Gram-negative bacteria. The enzymatic activity of OMPLA is strictly regulated to prevent uncontrolled breakdown of the surrounding phospholipids. The activity of OMPLA can be induced by membrane perturbation and concurs with dimerization of the enzyme. The recently elucidated crystal structures of the inactive, monomeric and an inhibited dimeric form of the enzyme provide detailed structural insight into the functional properties of the enzyme. OMPLA is a serine hydrolase with a unique Asn-156,His-142,Ser-144 catalytic triad. Only in the dimeric state, complete substrate binding pockets and functional oxyanion holes are formed. A model is proposed for the activation of OMPLA in which membrane perturbation causes the formation of non-bilayer structures, resulting in the presentation of phospholipids to the active site of OMPLA and leading to the formation of the active dimeric species. Possible roles for OMPLA in maintaining the cell envelope integrity and in pathogenicity are discussed. [source] | ||||||||||