4-Nitro-L-histidine as a substrate for histidine ammonia-lyase: the role of beta-hydrogen acidity in the rate-limiting step.

The K_m for the interaction of 4-nitro-L-histidine with histidine ammonia-lyase (reduced enzyme, pH 8.0) is comparable to that for L-histidine, while V_max is $\text{1}\text{8}$ that for the natural substrate. With the analog, addition of Cd⁺² effects a small decrease in K_m but fails to alter V_max; the normal deuterium isotope effect for removal of the β-hydrogen (1.5–2.0) is eliminated; and enzyme-catalyzed incorporation of solvent tritium into substrate occurs to a much greater extent than into histidine. Thus, the nitro group increases the acidity of the β-hydrogen and the stability of the conjugate carbanion to such a degree that CH bond cleavage now precedes rate-limiting CN bond cleavage.