Bioinformatic characterization of the extracellular lipases from Kluyveromyces marxianus

Abstract

Lipases are hydrolytic enzymes that break the ester bonds of triglycerides, generating free fatty acids and glycerol. Extracellular lipase activity has been reported for the nonconventional yeast Kluyveromyces marxianus, grown in olive oil as a substrate, and the presence of at least eight putative lipases has been detected in its genome.However, to date, there is no experimental evidence on the physiological role of the putative lipases nor their structural and catalytic properties. In this study, a bioinformatic analysis of the genes of the putative lipases from K. marxianus L-2029 was performed, particularly identifying and characterizing the extracellular expected enzymes, due to their biotechnological relevance. The amino acid sequence of 10 putative lipases, obtained by in silico translation, ranged between 389 and 773 amino acids. Two of the analysed putative proteins showed a signal peptide, 25 and 33 amino acids long for KmYJR107Wp and KmLIP3p, and a molecular weight of 44.53 and 58.23 kDa, respectively. The amino acid alignment of KmLIP3p and KmYJR107Wp with the crystallized lipases from a patatin and the YlLip2 lipase from Yarrowia lipolytica, respectively, revealed the presence of the hydrolase characteristic motifs. From the 3D models of putative extracellular K. marxianus L-2029 lipases, the conserved pentapeptide of each was determined, being GTSMG for KmLIP3p and GHSLG for KmYJR107Wp; besides, the genes of these two enzymes (LIP3 and YJR107W) are apparently regulated by oleate response elements. The phylogenetic analysis of all K. marxianus lipases revealed evolutionary affinities with lipases from abH15.03, abH23.01, and abH23.02 families. K E Y W O R D S bioinformatic tools, Kluyveromyces marxianus, lipases, phylogeny

Lipase activity of recombinant KmYJR107Wp and KmLIP3p enzymes expressed in Saccharomyces cerevisiae BY4742 from Kluyveromyces marxianus L2029

“…Considering the sequence of the PCR-amplified products and their in silico translation, the presence of signal peptide from both putative lipases was verified, as previously reported. 12 The nucleotide sequences of the promoter region and genes of both extracellular lipases of K. marxianus L-2029 are available in the NCBI database with accession numbers MH512931 for LIP3 and MH512932 for YJR107W.…”

“…After a bioinformatic analysis and a phylogenetic comparison with well-characterized fungal lipases of the amino acid putative lipases sequence, two of them, KmLIP3p and KmYJR107Wp, were identified as those likely responsible for K. marxianus extracellular lipase activity since both have a signal peptide and a high phylogenetic relationship with extracellular lipases. 12 However, the biotechnological potential of K. marxianus extracellular lipases is unknown.…”

Bioprospecting Kluyveromyces marxianus as a Robust Host for Industrial Biotechnology

“…From the 3D models of K. marxianus putative L-2029 lipases, the conserved pentapeptide was determined to be GHSLG for KmYJR107Wp and GTSMG for KmLIP3p . The phylogenetic analysis of all K. marxianus lipases demonstrated evolutionary affinities with lipases from abH23.01, abH23.02, and abH15.03 and families ( Martínez-Corona et al, 2020 ). In fact, microbial lipases are an interesting class of biocatalysts with prospective applications in medical and pharmaceutical, detergent, food, baking, dairy, flavor, organic synthesis, paper manufacturing, perfumery, cosmetics, leather, biosurfactant, agrochemical, oleochemical, bioremediation, and biosensors related industries ( Sarmah et al, 2018 ).…”