A new transaminase (VbTA) was identified from the genome of the halotolerant marine bacterium Virgibacillus 21D. Following heterologous expression in Escherichia coli, it was located entirely in the insoluble fraction. After a single mutation, identified via sequence homology analyses, the VbTA T16F mutant was successfully expressed in soluble form and characterised. VbTA T16F showed high stability towards polar organic solvents and salt exposure, accepting mainly hydrophobic aromatic amine and carbonyl substrates. The 2.0 Å resolution crystal structure of VbTA T16F is here reported, and together with computational calculations, revealed that this mutation is crucial for correct dimerisation and thus correct folding, leading to soluble protein expression.

Strategic single point mutation yields a solvent- and salt-stable transaminase from Virgibacillus sp. in soluble form / B. Guidi, M. Planchestainer, M.L. Contente, T. Laurenzi, I. Eberini, L.J. Gourlay, D. Romano, F. Paradisi, F. Molinari. - In: SCIENTIFIC REPORTS. - ISSN 2045-2322. - 8:1(2018 Nov 06). [10.1038/s41598-018-34434-3]

Strategic single point mutation yields a solvent- and salt-stable transaminase from Virgibacillus sp. in soluble form

B. Guidi
Primo
;
M.L. Contente;T. Laurenzi;I. Eberini;L.J. Gourlay;D. Romano;F. Molinari
Ultimo
2018

Abstract

A new transaminase (VbTA) was identified from the genome of the halotolerant marine bacterium Virgibacillus 21D. Following heterologous expression in Escherichia coli, it was located entirely in the insoluble fraction. After a single mutation, identified via sequence homology analyses, the VbTA T16F mutant was successfully expressed in soluble form and characterised. VbTA T16F showed high stability towards polar organic solvents and salt exposure, accepting mainly hydrophobic aromatic amine and carbonyl substrates. The 2.0 Å resolution crystal structure of VbTA T16F is here reported, and together with computational calculations, revealed that this mutation is crucial for correct dimerisation and thus correct folding, leading to soluble protein expression.
Settore BIO/10 - Biochimica
Settore AGR/13 - Chimica Agraria
   PIANO DI SOSTEGNO ALLA RICERCA 2015-2017 - LINEA 2 "DOTAZIONE ANNUALE PER ATTIVITA' ISTITUZIONALE"
6-nov-2018
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/601535
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