Identification of KASIII enzymes with novel substrate specificities: demonstration of in vivo production of novel ω‐1 hydroxylated fatty acids using a novel KASIII (967.7)

Most bacteria and plants typically synthesize straight and/or branched chain fatty acids. The proportion of these two classes of fatty acids is determined by availability of acyl‐CoA precursor pools and by substrate specificity of the enzyme that initiates fatty acid synthesis, 3‐Ketoacyl ACP Synthase III (KASIII). Presence of unusual fatty acids in certain organisms, such as ω‐cyclic or ω‐phenylic fatty acids, suggests that KASIII in such organisms have specificities for unusual acyl‐CoA substrates, such as cyclic or aromatic acyl‐CoAs. In this study, we have biochemically and biophysically characterized unique KASIII enzymes from four diverse bacterial sources, namely C. gingivalis , B. subtilis , T. aquaticus and A. acidocaldarius . From this set, we have identified several KASIIIs that possess novel substrate specificities towards atypical fatty acid synthesis primers, including hydroxylated (3‐hydroxybutyryl‐CoA), unsaturated (crotonyl‐CoA) and aromatic (benzoyl‐CoA) primers. Additionally, we have demonstrated in‐vivo production of ω‐1 hydroxy‐fatty acids in a modified strain of R. rubrum , by recombinantly co‐expressing A. acidocaldarius KASIII which utilizes 3‐hydroxybutyryl‐CoA. Identification of such unique KASIII enzymes introduces the opportunity to synthesize diverse ω‐functionalized fatty acids which can serve as chemical intermediates for production of bio‐based chemicals. Grant Funding Source : NSF