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. 2008 Nov 21;283(47):32328–32333. doi: 10.1074/jbc.M804332200

FIGURE 1.

FIGURE 1.

A, calorimetric profile of UDP-GlcA binding to K4CP. The reaction cell contained a solution of K4CP (1. 4 ml and 124.8 μm) in 25 mm HEPES buffer (pH 7.5) containing 20 mm MnCl2, 0.1 m NaCl, and 1 mm CaCl2. The syringe contained 1 mm UDP-GlcA in the same buffer. Top panel, raw calorimetric data obtained from the injection of 3 μl aliquots of UDP-GlcA at 3-min intervals. The lower panel shows the integrated binding isotherm with the experimental points (▪) and best fit. B, calorimetric profile of UDP-GalNAc binding to K4CP. C, calorimetric profile of UDP-GlcA binding to the K4CP mutant construct 239ACA241. D, gel showing the purity of K4CP used in ITC experiments and enzyme assays. The first lane contains the SeeBlue Plus 2 prestained molecular marker (Invitrogen), and the second lane contains purified K4CP.