Structures of N-acetylornithine transcarbamoylase from Xanthomonas campestris complexed with substrates and substrate analogs imply mechanisms for substrate binding and catalysis.

N-acetyl-L-ornithine transcarbamoylase (AOTCase) is a new member of the transcarbamoylase superfamily that is essential for arginine biosynthesis in several eubacteria. We report here crystal structures of the binary complexes of AOTCase with its substrates, carbamoyl phosphate (CP) or N-acetyl-L-ornithine (AORN), and the ternary complex with CP and N-acetyl-L-norvaline. Comparison of these structures demonstrates that the substrate-binding mechanism of this novel transcarbamoylase is different from those of aspartate and ornithine transcarbamoylases, both of which show ordered substrate binding with large domain movements. CP and AORN bind to AOTCase independently, and the main conformational change upon substrate binding is ordering of the 80's loop, with a small domain closure around the active site and little movement of the 240's loop. The structures of the complexes provide insight into the mode of substrate binding and the mechanism of the transcarbamoylation reaction.