Abstract
Cordycepin is an important nucleoside analogue and the principal active compound found in Cordyceps militaris. It exhibits anticancer, antiviral, immunomodulatory, and various other biological activities. However, the low cordycepin yields and high manufacturing costs limit its large-scale application. To develop a low-cost nutrient regulator to improve cordycepin production, we investigated the effect of corn steep liquor protein hydrolysate (CSLPH) on cordycepin biosynthesis during submerged fermentation of C. militaris, and the response mechanism of C. militaris to CSLPH utilization was revealed by comparative analysis of cell growth, cordycepin production, and transcriptomes. The results showed that a low CSLPH level in the medium stimulated the biosynthesis of cordycepin, and the addition of 1.0-3.5 g/L CSLPH in the medium increased the cordycepin production by 192.46% to 405.32%, which achieved the maximum cordycepin yield at a concentration of 1.5 g/L, and significantly promoted the growth of C. militaris after 14 days of fermentation. Transcriptomic analysis revealed that a set of significantly up-regulated genes in the pathway of cordycepin biosynthesis were enriched under CSL and CSLPH treatments, and some key up-regulated genes in the metabolic pathway of cordycepin biosynthesis were validated by quantitative real-time PCR. Moreover, transcriptomic analysis confirmed that the energy metabolism, amino acid metabolism, and the nucleotide pathway of Cordyceps militaris were improved by CSLPH, which facilitates cordycepin biosynthesis. Additionally, a cooperative mechanism by which CSLPH regulates cordycepin biosynthesis was elucidated for the first time in the current study. These results suggest that enzymatically hydrolyzed corn steep liquor is a more suitable alternative nitrogen source than corn steep liquor for efficient cordycepin production at low cost.