Discovery and engineering of an aldehyde tolerant 2-deoxy-D-ribose-5-phosphate aldolase (DERA) from Pectobacterium atrosepticum

MEERA HARIDAS; CAROLIN BISTERFELD; LE MIN CHEN; STEFAN R. MARSDEN; FABIO TONIN; ROSARIO MÉDICI; ADOLFO IRIBARREN; ELIZABETH LEWKOWICZ; PETER-LEON HAGEDOORN; ULF HANEFELD; EMAN ABDELRAHEEM

Catalysts

MPDI

Lugar: Basel; Año: 2020 vol. 10 p. 1 - 10

2073-4344

2-Deoxy-D-ribose 5-phosphate aldolase (DERA) is the only known aldolase that accepts two aldehyde substrates, which makes it an attractive catalyst to produce a chiral polyol motif which is required for certain pharmaceutical compounds (e.g. atorvastatin and pravastatin). However, inactivation of the enzyme in the presence of aldehydes hinders its practical application. Whole cells of Pectobacterium atrosepticum were reported to exhibit good acetaldehyde tolerance and catalyzed the aldol reaction to produce 2-deoxyribose 5-phosphate from two aldehyde molecules with good yields. The gene encoding P. atrosepticum (PaDERA) was identified and the enzyme was heterologously expressed in E.coli, as well as several site-directed mutants of the enzyme. The purification protocol was optimized and an initial characterization was conducted for the different mutants. Unlike other DERAs which show a maximal activity between pH 4.0 to 7.5, PaDERA presented an optimum working pH in the alkaline range between 8.0 and 9.0, which could warrant its use for specific syntheses in the future. PaDERA also displayed a 4-fold higher specific activity than the E. coli DERA (EcDERA) and displayed a promising acetaldehyde resistance outside the whole-cell environment. The C49M mutation, which was identified previously to increase acetaldehyde tolerance in EcDERA, also led to significant improvement in the acetaldehyde tolerance of PaDERA