TABLE 1

Effects of mutations in TrzN on protein yield, stability, and kinetic parametersa

Amino acidsbYield (mg/liter)Tm50(app) (oC) (holo-TrzN)kcat (s−1)Km (μM)kcat/Km (105 M−1 s−1)
D, L, A<0.252.12.119.71.1
N, L, A<0.1ND2.621.31.2
D, P, A0.651.33.148.60.6
D, L, V8.451.82.818.91.5
N, P, A1.252.33.252.20.6
N, L, V14.454.42.621.71.2
D, P, V28.251.03.254.20.6
N, P, V67.253.63.753.00.7
  • a Yield was measured as purified protein, stability was measured as the temperature at which 50% activity was lost (68), and kinetic parameters were measured with atrazine as the substrate using a previously published protocol (38). The Tm50(app) values could not be determined for the majority of the apoenzyme samples because the protein immediately unfolded upon removal of the metal ion.

  • b Amino acids at positions 38, 131, and 159, respectively.