Expanding the horizon of biodiesel production via enzyme engineering

ABSTRACT

Biodiesel generated from oils and fats composed of fatty acid alkyl esters is a potential substitution for fossil fuels, gaining attention as the demand for sustainable energy grows. Enzymatic transesterification has emerged as a promising method for biodiesel production due to the low energy required, easy glycerol recovery and compatibility with various feedstocks. This method also offers high specificity and operates under mild reaction conditions. However, limitations such as enzyme production cost, low product yield, and low stability under harsh conditions, have hindered the progress of biocatalysis. Protein engineering has emerged as an important strategy to overcome these limitations, particularly through rational design. Rational design approaches are advantageous in lipase engineering, due to accessible experimental tools and existing knowledge of lipase structure and function. Site-directed mutagenesis has been applied to improve stability, substrate specificity, methanol tolerance, and catalytic efficiency of lipase by introducing molecular interactions like hydrogen bonds and disulfide bridges as well as improved lid flexibility by mutations at the lid or lid swapping. This targeted approach allows for the precise modification of amino acid residues, optimizing lipase for better performance, which is crucial for efficient biodiesel production.

KEYWORDS:

• Biodiesel
• biocatalyst
• Lipase
• protein engineering
• transesterification

Abbreviations

NaOH:	=	Sodium hydroxide
KOH:	=	Potassium hydroxide
HCl:	=	Hydrochloric acid
H2SO4:	=	Sulfuric acid
CH3Ona:	=	Sodium methoxide
FFA:	=	Free fatty acid
FAME:	=	Methyl fatty acid esters
CALA:	=	Candida antarctica lipase
TLL:	=	Thermomyces lanuginosus lipase
pENO:	=	Enolase promoter
MRL:	=	Magnetic responsive lipase
FAME:	=	Fatty acid methyl esters
DAG:	=	Diacylglycerol
TAG:	=	Triacylglycerol
MAG:	=	Monoacylglycerol
WCO:	=	Waste cooking oil
%wt:	=	Weight percent

Acknowledgements

The authors express their gratitude for the financial support from the Research Management CentrePutra grant (GP berimpak project number 9708100), Universiti Putra Malaysia (Putra Grant).

Disclosure statement

No potential conflict of interest was reported by the author(s).

Authors contribution

Conceptualization, R.N.Z.R.A; M.S.M.A.; N.H.A.K.; writing—original draft preparation, S.N.H.I.; writing—review and editing, R.N.Z.R.A; M.S.M.A.; N.H.A.K.; A.L.T.C.; visualization, S.N.H.I.; All authors have read and agreed to the published version of the manuscript.

Additional information

Funding

The work was supported by the Universiti Putra Malaysia [9708100].