Skip to main content
. 2003 Dec;185(23):6764–6772. doi: 10.1128/JB.185.23.6764-6772.2003

FIG. 3.

FIG. 3.

Phosphorylation of His6-IIALacS by His6-HPr(Ser-P)(His∼P) and by His10-IIABLMan(His∼P) and His10-IIABHMan(His∼P). (A) The synthesis of His6-HPr(Ser-P)(His∼32P) is described in Materials and Methods. Phosphorylation of 5.8 μM His6-IIALacS by ca. 20 μM His6-HPr(Ser-P)(His∼32P) was conducted at 10°C. Samples were withdrawn at intervals, proteins were separated by SDS-PAGE, and phosphoproteins were revealed by autoradiography. (B) His10-IIABLMan(His∼32P) and His10-IIABHMan(His∼32P) were synthesized and purified as described in Materials and Methods. Phosphorylation of His6-IIALacS by His10-IIABLMan(His∼32P) and His10-IIABHMan(His∼32P) was carried out in 10 mM HEPES (pH 7.5), containing 5 mM MgCl2, either 2.8 μM His10-IIABLMan(His∼32P) or His10-IIABHMan(His∼32P), and 5.8 μM His6-IIALacS in a total volume of 30 μl. The proteins were separated by SDS-PAGE and revealed by autoradiography. Lanes: 1, phosphorylated products resulting from the incubation of His10-IIABHMan(His∼32P) with His6-HPr; 2, phosphorylated products resulting from the incubation of His10-IIABHMan(His∼32P) with His6-IIALacS; 3, phosphorylated products resulting from the incubation of His10-IIABLMan(His∼32P) with His6-IIALacS.