The alpha-amylase MrAMY1 is better than MrAMY2 in rice starch degradation, which promotes Monascus pigments production in Monascus ruber.

Previously, AoamyA, the alpha amylase-encoding gene from Aspergillus oryzae, was heterologously expressed in Monascus ruber CICC41233 to promote starch hydrolysis and increase the production of Monascus pigments. The target of this study is to screen the effective alpha-amylases from M. ruber for starch fast degradation and investigated for Monascus pigments production. The 13 types of predicted alpha-amylases in the M. ruber NRRL1597 genome were divided into four classes based on EC number and into five groups based on the glycoside hydrolase sub-family. The predicted alpha-amylases MrAMY1 (protein ID 440333) and MrAMY2 (protein ID 324551) showed the closest match with AOamyA by phylogenetic analysis. The genes encoding alpha-amylase, Mramy1and Mramy2, were cloned from M. ruber CICC41233. However, the gene sequence of Mramy1 from M. ruber CICC41233 differed from that of M.ruber NRRL1597 in the length of the intron sequence. Furthermore, the Mramy1-overexpressed strain M.ruber 440333-6A completely degraded the starch of rice grain in 2 d; in contrast, starch (40.32 mg/mL) remained when rice grain was incubated with the Mramy2-overexpressed strain, M. ruber 324551-D even after 2 d, while 45.43 mg/mL and 10.48 mg/mL starch remained after 2 d and 6 d, respectively, in wild type M. ruber CICC41233. Compared to that of M. ruber CICC41233, the total Monascus pigments and ethanol-soluble pigments in M.ruber 440333-6A increased by 71.69% and 119.33% after 6d, respectively; however, it decreased by 21.40%and 26.58% after 6d, respectively, in M. ruber 324551-D. This study demonstrated that alpha-amylase MrAMY1 was superior to MrAMY2, as it effectively degraded the starch of rice grain and enhanced Monascus pigments production. © King Abdulaziz City for Science and Technology 2020.