Regulation of Riboflavin Biosynthesis in Microorganisms and Construction of the Advanced Overproducers of This Vitamin.

Riboflavin (vitamin B2) is an essential micronutrient required for all living organisms. It is naturally synthesized by plants and most microorganisms, including the bacterium Bacillus subtilis, the filamentous fungus Ashbya gossypii, and the yeast Candida famata-all of which are known to be riboflavin overproducers. The choice of production organism in industrial applications depends on factors such as yield, ease of cultivation, and the availability of genetic tools. As a result, several microorganisms are commonly used, and their relative prominence can shift over time with advances in metabolic engineering and process optimization. This review presents a comparative analysis of riboflavin biosynthesis across prokaryotic and eukaryotic systems, with a particular focus on regulatory mechanisms governing flavinogenesis. Special attention is given to recent advances in metabolic engineering strategies, including the application of CRISPR/Cas9 genome editing in Bacillus subtilis and Ashbya gossypii. In yeast systems, significant improvements in riboflavin production have been achieved primarily through the manipulation of transcriptional regulators (e.g., SEF1, SFU1, TUP1) and metabolic genes. The role of other important genes (PRS3, ADE4, ZWF1, GND1, RFE1, VMA1, etc.) in riboflavin overproduction in C. famata is described. The review also explores the use of alternative, low-cost feedstocks-including lignocellulosic hydrolysates and dairy by-products-to support more sustainable and economically viable riboflavin production. Although considerable progress has been achieved in genetic optimization and bioprocess development, further work is required to fine-tune metabolic flux and maximize riboflavin synthesis, particularly under industrial conditions. This review highlights key opportunities for future research aimed at refining metabolic interventions and expanding the use of renewable substrates for environmentally sustainable riboflavin production.