Identification of Thermoregulatory Factors Involved in Validamycin Biosynthesis and Reconstitution of a Thermoregulated Expression System

ABSTRACT Thermoregulatory systems are rarely utilized in the synthetic biology of Streptomyces . In Streptomyces hygroscopicus TL01, the previous finding of enhanced validamycin biosynthesis at 37°C indicated the involvement of a thermoregulatory system. In this study, the P valA ( P KA ) promoter was thermoregulated in Streptomyces albus J1074, and its regulation was determined by a GC‐rich DNA clamp. P KA ‐binding regulatory proteins were identified through DNA affinity chromatography. Overexpression of SHJG_5852 and SHJG_8253 in S. albus resulted in the P KA strength ratios of 2.61 and 4.34 at 37°C and 30°C, respectively. In contrast, the overexpression of SHJG_6392 led to an inhibited transcription of the P KA at 30°C. A thermoregulatory system was reconstituted in S. albus J1074 with a shorter but more efficient promoter derived from the P KA , SHJG_5852 , and SHJG_6392 . With this system, the induced indigoidine titer at 37°C was 3.56 times that at 30°C. In summary, we identified thermoregulatory factors involved in validamycin biosynthesis and reconstituted a system for the thermoregulated production of actinobacterial natural products.