Expanding the repertoire of nitrilases with broad substrate specificity and high substrate tolerance for biocatalytic applications

2020 ◽  
Vol 94 ◽  
pp. 289-296 ◽  
Author(s):  
Avinash Vellore Sunder ◽  
Shikha Shah ◽  
Pratima Rayavarapu ◽  
Pramod P. Wangikar
Keyword(s):  
Substrate Specificity ◽  
Broad Substrate Specificity ◽  
High Substrate ◽  
Substrate Tolerance

scholarly journals Kinetic Crystallography on MutT and its homolog

2014 ◽  
Vol 70 (a1) ◽  
pp. C486-C486
Author(s):  
Teruya Nakamura ◽  
Keisuke Hirata ◽  
Kohei Yoshikawa ◽  
Miyuki Inazato ◽  
Mami Chirifu ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Activation Mechanism ◽  
Structural Basis ◽  
Human Homolog ◽  
Substrate Specificities ◽  
Broad Substrate Specificity ◽  
E Coli ◽  
High Substrate ◽  
Substrate Complex ◽  
Complex Crystals

Oxidized deoxynucleotides cause replicational errors because of their misincorporations into DNA. The MutT and related proteins prevent transversion mutations by hydrolyzing mutagenic oxidized nucleotides such as 8-oxo-dGTP and 2-oxo-dATP, and there is a difference in substrate specificities between them. E. coli MutT hydrolyzes 8-oxo-dGTP to 8-oxo-dGMP with extremely high substrate specificity. On the other hand, its human homolog has broad substrate specificity for oxidized nucleotides and hydrolyzes 8-oxo-dGTP as well as 2-oxo-dATP. In order to understand mechanisms of their substrate specificities, we solved the crystal structures of MutT and its homolog complexed with their substrates and revealed structural basis of the high substrate specificity of E. coli MutT for 8-oxoguanine nucleotide and the broad substrate specificity of its human honolog for oxidized nucleotides. In this paper, we report the hydrolysis mechanisms of both enzymes revealed by kinetic protein crystallography. Both hydrolysis reactions were initiated by soaking the enzyme-substrate complex crystals in divalent metal solution. After incubation under various conditions, the reactions were terminated by freezing the crystals at 100K. X-ray diffraction data were collected at Spring-8 and Photon Factory. In the MutT crystals, the structures of sequential catalytic intermediates showed the activation mechanism of the nucleophilic water molecule synchronized with the coordination of metal ions. Now by using the crystals of its human homolog, the trial of the catching the intermediate state of catalysis is in progress.

scholarly journals LASS3 (longevity assurance homologue 3) is a mainly testis-specific (dihydro)ceramide synthase with relatively broad substrate specificity

2006 ◽  
Vol 398 (3) ◽  
pp. 531-538 ◽  
Author(s):  
Yukiko Mizutani ◽  
Akio Kihara ◽  
Yasuyuki Igarashi
Keyword(s):  
Amino Acid ◽  
Substrate Specificity ◽  
Family Members ◽  
Fatty Acyl ◽  
Ceramide Synthase ◽  
Broad Substrate Specificity ◽  
Acid Protein ◽  
Amino Acid Protein

The LASS (longevity assurance homologue) family members are highly conserved from yeasts to mammals. Five mouse and human LASS family members, namely LASS1, LASS2, LASS4, LASS5 and LASS6, have been identified and characterized. In the present study we cloned two transcriptional variants of hitherto-uncharacterized mouse LASS3 cDNA, which encode a 384-amino-acid protein (LASS3) and a 419-amino-acid protein (LASS3-long). In vivo, [3H]dihydrosphingosine labelling and electrospray-ionization MS revealed that overproduction of either LASS3 isoform results in increases in several ceramide species, with some preference toward those having middle- to long-chain-fatty acyl-CoAs. A similar substrate preference was observed in an in vitro (dihydro)ceramide synthase assay. These results indicate that LASS3 possesses (dihydro)ceramide synthesis activity with relatively broad substrate specificity. We also found that, except for a weak display in skin, LASS3 mRNA expression is limited almost solely to testis, implying that LASS3 plays an important role in this gland.

scholarly journals TheArabidopsisAtSTE24 Is a CAAXProtease with Broad Substrate Specificity

2002 ◽  
Vol 277 (33) ◽  
pp. 29856-29864 ◽  
Author(s):  
Keren Bracha ◽  
Meirav Lavy ◽  
Shaul Yalovsky
Keyword(s):  
Substrate Specificity ◽  
Broad Substrate Specificity

scholarly journals A new dopachrome-rearranging enzyme from the ejected ink of the cuttlefish Sepia officinalis.

1994 ◽  
Vol 299 (3) ◽  
pp. 839-844 ◽  
Author(s):  
A Palumbo ◽  
M d'Ischia ◽  
G Misuraca ◽  
L De Martino ◽  
G Prota
Keyword(s):  
Catalytic Activity ◽  
Methyl Ester ◽  
Substrate Specificity ◽  
Molecular Mass ◽  
Significant Inhibition ◽  
Single Band ◽  
Pigment Cells ◽  
Sepia Officinalis ◽  
High Substrate ◽  
Sds Page

A melanogenic enzyme catalysing the rearrangement of dopachrome has been identified in the ejected ink of the cuttlefish Sepia officinalis. This enzyme occurs as a heat-labile protein which co-migrates with tyrosinase under a variety of chromatographic and electrophoretic conditions. On SDS/PAGE it shows like a single band with an approx. molecular mass of 85 kDa. The enzyme possesses high substrate specificity, acting on L-dopachrome (Km = 1 mM at pH 6.8) and on L-alpha-methyl-dopachrome, but not on D-dopachrome, L-dopachrome methyl ester, dopaminochrome and adrenochrome. Significant inhibition of the catalytic activity was observed with tropolone and L-mimosine. H.p.1.c. analysis of the enzyme-catalysed rearrangement of L-dopachrome revealed the quantitative formation of the decarboxylated product, 5,6-dihydroxyindole. These results point to marked differences between melanogenesis in cephalopod pigment cells and in melanocytes, which may have important implications in relation to the use of sepiomelanin as a model for studies of mammalian melanins.

Prediction of amino acid residues participated in substrate recognition by cytochrome P450 subfamilies with broad substrate specificity

2013 ◽  
Vol 26 (2) ◽  
pp. 86-91 ◽  
Author(s):  
Maria S. Zharkova ◽  
Boris N. Sobolev ◽  
Nina Yu. Oparina ◽  
Alexander V. Veselovsky ◽  
Alexander I. Archakov
Keyword(s):  
Cytochrome P450 ◽  
Amino Acid ◽  
Substrate Specificity ◽  
Substrate Recognition ◽  
Amino Acid Residues ◽  
Broad Substrate Specificity
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