Figure 2. The active site of Na⁺-free wild-type thrombin is non-catalytic.

(A) The stereo representation of electron density (blue) surrounding residues 57, 191–195 and 214–221 contoured at 2σ indicates the high quality of the structure in the active-site region. Carbon atoms are shown in yellow, oxygen in red, nitrogen in blue, sulphur in green, and waters are magenta. (B) The hydrogen-bonding interactions in the active site are extensive and non-catalytic (green broken lines). Of particular importance for the activity of thrombin is the movement of Glu¹⁹², which results in a main-chain hydrogen bond to the side-chain Hγ and main-chain N–H of Ser¹⁹⁵, and the flipping of the oxyanion hole partner Gly¹⁹³. The atoms which normally constitute the oxyanion hole (Gly¹⁹³ and Ser¹⁹⁵ nitrogens) are indicated by arrows. (C) A superposition of the same active-site residues for Na⁺-free wild-type thrombin (yellow), recombinant slow thrombin E217K variant (cyan) and normal fast thrombin (magenta) illustrates the radical nature of the active site perturbations caused by the absence of Na⁺. The active-site geometry is similarly perturbed for the E217K variant, but the perpendicular orientation of the side chain of Trp²¹⁵ further blocks the active site. The orientation of the Trp²¹⁵ side chain in Na⁺-free thrombin is parallel to that of fast thrombin, but has flipped 180° to form a hydrogen bond with Glu²¹⁷ (see B).