A novel cold-adapted lipase from Acinetobacter sp. XMZ-26: gene cloning and characterisation

Abstract

Acinetobacter sp. XMZ-26 (ACCC 05422) was isolated from soil samples obtained from glaciers in Xinjiang Province, China. The partial nucleotide sequence of a lipase gene was obtained by touchdown PCR using degenerate primers designed based on the conserved domains of cold-adapted lipases. Subsequently, a complete gene sequence encoding a 317 amino acid polypeptide was identified. Our novel lipase gene, lipA, was overexpressed in Escherichia coli. The recombinant protein (LipA) was purified by Ni-affinity chromatography, and then deeply characterised. The LipA resulted to hydrolyse pNP esters of fatty acids with acyl chain length from C2 to C16, and the preferred substrate was pNP octanoate showing a k(cat) = 560.52 ± 28.32 s(-1), K(m) = 0.075 ± 0.008 mM, and a k(cat)/K(m) = 7,377.29 ± 118.88 s(-1) mM(-1). Maximal LipA activity was observed at a temperature of 15°C and pH 10.0 using pNP decanoate as substrate. That LipA peaked at such a low temperature and remained most activity between 5°C and 35°C indicated that it was a cold-adapted enzyme. Remarkably, this lipase retained much of its activity in the presence of commercial detergents and organic solvents, including Ninol, Triton X-100, methanol, PEG-600, and DMSO. This cold-adapted lipase may find applications in the detergent industry and organic synthesis.

Expression, purification and characterization of a functional, recombinant, cold-active lipase (LipA) from psychrotrophic Yersinia enterocolitica

“…LipA retained activity at pH 5.0-11.0, with its maximum activity at pH 7.5, whereas it was reported that most cold-active lipases had optimal activities at neutral or alkaline pH [18][19][20][21][22]. The optimum temperature of the purified lipase was found to be 25°C, which is consistent with that of other cold-active lipases, such as Microbacterium phyllosphaerae MTCC 7530 [23], Photobacterium lipolyticum M37 [24], Pseudoalteromonas sp.…”

An Organic Solvent-Tolerant Alkaline Lipase from Cold-Adapted <i>Pseudomonas mandelii</i>: Cloning, Expression, and Characterization

“…[7][8][9] Here, we report the cloning, expression, and characterization of an organic solvent-tolerant alkaline lipase, LipS (GenBank ID JQ071496), from Pseudomonas mandelii JR-1. P. mandelii is a cold-adapted bacterium that grows at 4 C but not at 37 C, with an apparent optimum temperature range of 25-30 C. 10,11) To produce a clone, we utilized the genome sequence of Pseudomonas fluorescens Pf0-1, 12) which is closely related to P. mandelii, from among completed microbial genomes.…”

Expression and Characterization of a Novel Enantioselective Lipase from Aspergillus fumigatus

“…It showed the maximum activity and stability at 15°C, similar to the characteristics of a cold-active lipase from Acinetobacter sp. XMZ-26 (15°C at pH 9.0) [30], but much different from the normal fungal lipases which are generally active and stable at 40-50°C [31]. Also, in contrast with previous reported majority lipases from Aspergillus with acidic pH optima (5.0-6.0) [20], AFL67 showed an optimum pH of 7.5 and was almost inactivated below pH 6.5.…”