Anti-GH1 monoclonal antibody, clone 2I2.2

Background: Rice Os9BGlu31 is a transglucosidase that can transfer glucose to phenolic acids, flavonoids, and phytohormones. Os9BGlu31 displays a broad specificity with phenolic 1-O-beta-D-glucose esters acting as better glucose donors than glucosides, whereas the free phenolic acids of these esters are also excellent acceptor substrates. Methods: Based on homology modeling of this enzyme, we made single point mutations of residues surrounding the acceptor binding region of the Os9BGlu31 active site. Products of the wild type and mutant enzymes in transglycosylation of phenolic acceptors from 4-nitrophenyl beta-D-glucopyranoside donor were identified and measured by UPLC and negative ion electrospray ionization tandem mass spectrometry (LCMSMS). Results: The most active variant produced was W243N, while I172T and L183Q mutations decreased the activity, and other mutations at W243 (A, D, M, N, F and Y) had variable effects, depending on the acceptor substrate. The Os9BGlu31 W243N mutant activity was higher than that of wild type on phenolic acids and kaempferol, a flavonol containing 4 hydroxyl groups, and the wild type Os9BGlu31 produced only a single product from each of these acceptors in significant amounts, while W243 variants produced multiple glucoconjugates. Fragmentation analysis provisionally identified the kaempferol transglycosylation products as kaempferol 3-O, 7-O, and 4'-O glucosides and 3,7-O, 4',7-O, and 3,4'-O bis-D-glucosides. The Os9BGlu31 W243 mutants were also better able to use kaempferol 3-D-glucoside as a donor substrate. General significance: The W243 residue was found to be critical to the substrate and product specificity of Os9BGlu31 transglucosidase and mutation of this residue allows production of a range of glucoconjugates. (C) 2015 Elsevier B.V. All rights reserved.

The somatotropic axis, the control system for growth hormone (GH) secretion and its endogenous factors involved in the regulation of metabolism and energy partitioning, has promising potentials for producing economically valuable traits in farm animals. Here we investigated single nucleotide polymorphisms (SNPs) of the genes of factors involved in the somatotropic axis for growth hormone (GH1), growth hormone receptor (GHR), ghrelin (GHRL), insulin-like growth factor 1 (IGF-I) and leptin (LEP), using polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) and DNA sequencing methods in 452 individual Mehraban sheep. A nonradioactive method to allow SSCP detection was used for genomic DNA and PCR amplification of six fragments: exons 4 and 5 of GH1; exon 10 of GH receptor (GHR); exon 1 of ghrelin (GHRL); exon 1 of insulin-like growth factor-I (IGF-I), and exon 3 of leptin (LEP). Polymorphisms were detected in five of the six PCR products. Two electrophoretic patterns were detected for GH1 exon 4. Five conformational patterns were detected for GH1 exon 5 and LEP exon 3, and three for IGF-I exon I. Only GHR and GHRL were monomorphic. Changes in protein structures due to variable SNPs were also analyzed. The results suggest that Mehraban sheep, a major breed that is important for the animal industry in Middle East countries, has high genetic variability, opening interesting prospects for future selection programs and preservation strategies. (C) 2013 Elsevier B.V. All rights reserved.