A newly isolated yeast as an expression host for recombinant lipase

Abstract

Pichia guilliermondii strain SO isolated from spoiled orange was developed for use as an alternative expression host by using Pichia pastoris as the model of the experiment. This is the first study to report on the capability of P. guilliermondii SO as a host to express thermostable T1 lipase from Geobacillus zalihae. Alcohol oxidase and formaldehyde dehydrogenase promoters were present in the yeast genome. Interestingly, the recombinant yeast [SO/pPICZαB/T1-2 (SO2)] took only 30 h to reach optimal production with minimal methanol induction [1.5% (v/v)] in YPTM medium, as compared to P. pastoris, which took longer to reach its optimal condition. The purification yield of the His-tagged fusion lipase was 68.58%, with specific activity of 194.58 U/mg. The optimum temperature was 65°C at pH 9 in glycine-NaOH buffer, and it was stable up to 70°C in a wide pH range from pH 5 to 12. In conclusion, a newly isolated yeast from spoiled orange has been proven suitable for use as an expression host.

Optimization of physical conditions for the production of thermostable T1 lipase in Pichia guilliermondii strain SO using response surface methodology

“…However, a new yeast P. guilliermondii isolated from spoilt orange was used as a recombinant host for the expression of thermostable T1 lipase gene isolated from Geobacillus zalihae [ 17 , 18 ]. Similar to methylotrophic yeasts, gene expression and protein production in P. guilliermondii have been found to be driven by methanol induced AOXp 1 [ 18 ].…”

“…However, a new yeast P. guilliermondii isolated from spoilt orange was used as a recombinant host for the expression of thermostable T1 lipase gene isolated from Geobacillus zalihae [ 17 , 18 ]. Similar to methylotrophic yeasts, gene expression and protein production in P. guilliermondii have been found to be driven by methanol induced AOXp 1 [ 18 ]. In this study, P. guilliermondii was found capable of generating biomass using glycerol as a carbon source and relatively undergoing gene expression without any carbon source as observed in the study on Hansenula polymorpha [ 10 ] depending on the level of de-repression.…”

“…Previously, Escherichia coli has been used as a recombinant host for T1 lipase production, but was characterised by low expression [ 19 ]. It is important to note that T1 lipase gene has been also cloned and expressed in P. pastoris but recently, it was observed that P. guilliermondii was observed as a better producer of T1 lipase under the control of AOXp 1 [ 18 ]. Therefore, it is appropriate to screen and optimise the enzyme production, initially by media optimisation, followed by optimisation of physical conditions significant for the enhancement of T1 lipase production in P. guilliermondii .…”

Diversity and occurrence of methylotrophic yeasts used in genetic engineering

“…Since the majority of methylotrophic yeasts have similar methanol-inducible promoter in methanol utilization paths (Hartner, Glieder, 2006), the research group from Malaysia tested the assumption that expression constructs developed on the basis of K. phaffii could be used for expression of recombinant proteins in other methylotrophic yeasts (Oslan et al, 2015). They isolated the strain called Pichia sp.…”

Molecular expression of a recombinant thermostable bacterial amylase from Geobacillus stearothermophilus SR74 using methanol-free Meyerozyma guilliermondii strain SO yeast system

“…Moreover, a newly developed expression host, M. guillermondii strain SO, has the potential to produce foreign protein at a faster rate than P. pastoris with minimal methanol induction (Oslan et al 2015). P. pastoris requires 60 h, whereas M. guillermondii strain SO requires 30 h to produce foreign protein at an optimum level (Oslan et al 2015).…”