Efficient Biocatalytic Synthesis of (R)‐Pantolactone

Abstract Screening for stereoselective cyanohydrin synthesis in 96‐well plates was employed in the development of an efficient, pH‐stable hydroxynitrile lyase for the conversion of sterically hindered aliphatic aldehydes. Site‐saturation mutagenesis (SSM) resulted in a powerful catalyst for the stereoselective conversion of hydroxypivalaldehyde and pivalaldehyde to their corresponding ( R )‐cyanohydrins ( ee >97%) which are used as chiral building blocks (e.g., for pantothenic acid production). Furthermore, redesigning the Pa HNL5 gene and improving its expression by Pichia pastoris with the help of a new P AOX1 promoter variant and the helper protein PDI (protein disulfide isomerase) led to elevated amounts of today’s most efficient biocatalyst for vitamin B 5 synthesis.