Heterologous expression and biochemical characterization of a cold-active lipase from Rhizopus microsporus suitable for oleate synthesis and bread making

Abstract

Objectives Cold-active lipases which show high specific activity at low temperatures are attractive in industrial applications in terms of product stability and energy saving. We aimed to identify novel cold-active lipase suitable for oleates synthesis and bread making. Results A novel lipase gene (RmLipA) from Rhizopus microsporus was cloned and heterologously expressed in Pichia pastoris. The encoding sequence displayed 75% identity to the lipase from R. niveus. The highest extracellular lipase activity of 7931 U/mL was achieved in a 5-L fermentation. The recombinant enzyme (RmLipA) was optimally active at pH 8.0 and 20-25 °C, respectively, and stable over a wide pH range of 2.0-11.0. The enzyme was a cold-active lipase, exhibiting [ 80% of its maximal activity at 0 °C. RmLipA was a sn-1,3 regioselective lipase, and preferred to hydrolyze pNP esters and triglycerides with relatively long chain fatty acids. RmLipA synthesized various oleates using oleic acid and different alcohols as substrates ([ 95%). Moreover, it significantly improved the quality of bread by increasing its specific volume (21.7%) and decreasing its crumb firmness (28.6%). Conclusions A novel cold-active lipase gene from R. microsporus was identified, and its application potentials were evaluated. RmLipA should be a potential candidate in oleates synthesis and bread making industries.

Characterization and Molecular Dynamics Simulation of a Lipase Capable of Improving the Functional Characteristics of an Egg-Yolk-Contaminated Liquid Egg White

“…The enzyme activity was measured in different pH buffers under the optimal temperature conditions mentioned above, to determine the optimal pH of enzymes [18]. The buffer solutions were configured as follows: citric-acid-phosphate buffer (pH = 6-7), Tris-HCl buffer (pH = 8-9), and sodium-carbonate-sodium-bicarbonate buffer (pH = 10).…”

Mitochondrial genomic characteristics and phylogenetic analysis of a brewing fungus, Rhizopus microsporus Tiegh. 1875 (Mucorales: Rhizopodaceae)

“… 2020 ; Xiang et al. 2021 ). These properties make R. microsporus a highly valuable resource for industrial and agricultural applications.…”

Cold-Active Lipases and Esterases: A Review on Recombinant Overexpression and Other Essential Issues

“…In recent years, there have been many articles published on cold-active enzymes [ 30 , 31 , 32 ]. Previous studies on cold-active lipases and esterases focused on their isolation from various sources, overexpression, and biochemical characterisation [ 33 , 34 , 35 , 36 ]. Still, their purification and three-dimensional (3D) structures have received little attention.…”