Cloning, expression, purification, crystallization and preliminary X-ray characterization of allantoinase from Bacillus licheniformis ATCC 14580

Allantoinase, a member of the amidohydrolase superfamily, exists in a wide variety of organisms, including bacteria, fungi, plants and a few animals, such as fishes and amphibians. Allantoinase catalyzes the reversible hydrolysis of allantoin into allantoate by hydrolytic cleavage of the N1—C2 amide...

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Published in:Acta crystallographica. Section F, Structural biology communications Structural biology communications, 2014-11, Vol.70 (11), p.1513-1516
Main Authors: Conejero-Muriel, Mayte, Martínez-Gómez, Ana Isabel, Martínez-Rodríguez, Sergio, Gavira, Jose A.
Format: Article
Language:English
Subjects:
allantoin
allantoinase
amidohydrolase
Amidohydrolases - biosynthesis
Amidohydrolases - chemistry
Amidohydrolases - isolation & purification
Bacillus - enzymology
Bacillus licheniformis
Cloning, Molecular - methods
Crystallization Communications
Gene Expression Regulation, Bacterial
X-Ray Diffraction
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Summary:Allantoinase, a member of the amidohydrolase superfamily, exists in a wide variety of organisms, including bacteria, fungi, plants and a few animals, such as fishes and amphibians. Allantoinase catalyzes the reversible hydrolysis of allantoin into allantoate by hydrolytic cleavage of the N1—C2 amide bond of the five‐membered hydantoin ring. Allantoinase from Bacillus licheniformis (AllBali) presents an inverted enantioselectivity towards allantoin (R‐enantioselective), which is a distinguishable feature that is not observed for other allantoinases. In this work, B. licheniformis ATCC 14580 allantoinase (AllBali) containing a C‐terminal His6 tag was overproduced in Escherichia coli and purified to homogeneity. Crystals of AllBali were obtained by the vapour‐diffusion method using 0.1 M potassium thiocyanate, 20%(w/v) polyethylene glycol 3350 as a crystallization solution. X‐ray diffraction data were collected to a resolution of 3.5 Å with an Rmerge of 29.2% from a crystal belonging to space group P1211, with unit‐cell parameters a = 54.93, b = 164.74, c = 106.89 Å, β = 98.49°. There are four molecules in the asymmetric unit with a solvent content of 47% as estimated from the Matthews coefficient (VM = 2.34 Å3 Da−1).
ISSN:2053-230X
2053-230X
DOI:10.1107/S2053230X14021608