Table 4.

In vitro properties of modified forms of NACa

ConstructApparent molecular size in solution (kDa)FormbGel mobility shiftcTranscription activationc
NACEI
MBP-NACE75 (66)M (D)Y (Y)(Y)
56his6-NACEY (Y)N (N)
NACE-his6I
NACE100-his6I
MBP-NACE100(Y)
NACK66DYY
MBP-NACK70 (66)M (D)Y (Y)N (Y)
56his6-NACK77DY
NACK-his6YY
MBP-NACK12956 (28)M (D)(Y)
NACK120-his627DYY
NACK100-his622DY
  • a NACE, NACK, and several modified forms thereof were purified as described in Materials and Methods. The apparent molecular size in solution was determined by gel filtration, and this size was used to infer whether the product was monomeric or dimeric in solution. The ability to bind DNA was determined in a gel mobility shift assay using the urease operon promoter (ureDp) as a target. The ability to activate transcription from the hutUH promoter (hutUp) was tested as described in the legend to Fig. 2. The values in parentheses were determined after cleavage with Factor Xa or thrombin.

  • b M, monomer; I, insoluble; D, dimer.

  • c Y, yes; N, no.