New insights into the binding mode of coenzymes: structure ofThermus thermophilusΔ1-pyrroline-5-carboxylate dehydrogenase complexed with NADP+

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Abstract
1-Pyrroline-5-carboxylate dehydrogenase (P5CDh) is known to preferentially use NAD+ as a coenzyme. The k cat value of Thermus thermophilus P5CDh (TtP5CDh) is four times lower for NADP+ than for NAD+. The crystal structure of NADP+-bound TtP5CDh was solved in order to study the structure-activity relationships for the coenzymes. The binding mode of NADP+ is essentially identical to that in the previously solved NAD+-bound form, except for the regions around the additional 2'-phosphate group of NADP+. The coenzyme-binding site can only accommodate this group by the rotation of a glutamate residue and subtle shifts in the main chain. The 2'-phosphate of NADP+ increases the number of hydrogen bonds between TtP5CDh and NADP+ compared with that between TtP5CDh and NAD+. Furthermore, the phosphate of the bound NADP+ would restrict the `bending' of the coenzyme because of steric hindrance. Such bending is important for dissociation of the coenzymes. These results provide a plausible explanation of the lower turnover rate of NADP+ compared with NAD+.