Table 2.

Glutamine-dependent CP synthesis activities for wild-type and mutant CPSs

	CP formation (μmoles • min−1 • mg−1)
	ATP	ATP + GTP
WT	0.64	0.81
A144Q	0.81	0.85
D207A	0.24	0.35
S209A	0.32	0.30
ADS	0.06*	0.07*
D207N	0.26	0.38
A144Q/D207A	0.37	0.40*
I211S	0.57	0.56
P690Q	0.71	0.78
D753A	0.06*	0.08
F755A	0.58	0.62
PDF	0.08	0.07
D753N	0.08	0.08

CP formation was measured by coupling to the reaction of ornithine transcarbamoylase in the presence of ornithine and determining the resulting citrulline. Assay mixtures included 50 mM HEPES, 100 mM KCl, 20 mM MgSO₄, 40 mM NaHCO₃, 10 mM glutamine, 1 mM DTT, 5 u/mL ornithine transcarbamoylase, 10 mM ornithine, and 5 mM each indicated nucleotide, final pH 7.6. CPS (5–10 μg) was incubated at 37°C for 5–60 min, depending on the activity of the particular mutant. Citrulline was determined by reaction with diacetyl monoxime (Guthohrlein and Knappe 1968). SEM for all values was less than 1%, except * = less than 5%.