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Title: Cloning, expression, and molecular dynamics simulations of a xylosidase obtained from Thermomyces lanuginosus
Authors: Gramany, Vashni 
Khan, Faez Iqbal 
Govender, Algasan 
Bisetty, Krishna 
Singh, Suren 
Permaul, Kugenthiren 
Keywords: Cloning;Expression;Structure prediction;Molecular dynamics
Issue Date: 19-Oct-2015
Publisher: Taylor and Francis Online
Source: Gramany, V. et al. 2015. Cloning, expression, and molecular dynamics simulations of a xylosidase obtained from Thermomyces lanuginosus. Journal of Biomolecular Structure and Dynamics. 34(8): 1681-1692.
Journal: Journal of biomolecular structure & dynamics 
The aim of this study was to clone, express, and characterize a β-xylosidase (Tlxyn1) from the thermophilic fungus Thermomyces lanuginosus SSBP in Pichia pastoris GS115 as well as analyze optimal activity and stability using computational and experimental methods. The enzyme was constitutively expressed using the GAP promoter and secreted into the medium due to the alpha-mating factor secretion signal present on the expression vector pBGPI. The 1276 bp gene consists of an open reading frame that does not contain introns. A 12% SDS–PAGE gel revealed a major protein band at an estimated molecular mass of 50 kDa which corresponded to zymogram analysis. The three-dimensional structure of β-xylosidase was predicted, and molecular dynamics simulations at different ranges of temperature and pH were performed in order to predict optimal activity and folding energy. The results suggested a strong conformational temperature and pH dependence. The recombinant enzyme exhibited optimal activity at pH 7 and 50°C and retained 80% activity at 50°C, pH 7 for about 45 min. This is the first report of the cloning, functional expression, and simulations study of a β-xylosidase from Thermomyces species in a fungal host.
ISSN: 0739-1102
Appears in Collections:Research Publications (Applied Sciences)

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