Bridging the Gap Between Theory and Experiment to Derive a Detailed Understanding of Hammerhead Ribozyme Catalysis - Progress in Molecular Biology and Translational Science - page 71

Table 2.10 Characterization of the active site structure and fluctuations for the G5
mutants in reactant states
WT U7C G5I G5A G5D

r Nu 4.07 (0.25) 3.78 (0.43) 4.18 (0.14) 3.84 (0.46) 3.94 (0.39)

y inl 124.3 (8.6) 133.3 (15.1) 118.3 (9.2) 129.8 (15.2) 128.1 (14.1)

F 0.25 (0.11) 0.38 (0.2) 0.2 (0.04) 0.35 (0.19) 0.31 (0.17)

d 0 4.64 (0.56) 4.95 (0.8) 5.2 (0.43)

3.97 (0.39) 4.27 (0.63)

r HB 2.07 (0.25) 2.1 (0.28) 2.22 (0.76) 2.08 (0.33) 2.31 (0.64)

y HB 152.7 (13.4) 153.6 (13.4) 155.5 (14.4) 156.9 (12.7) 149.5 (18)

% 98 97 91 97 84

% 62 64 72 75 56

r HA 2.75 (0.45) 3.41 (1.02) 5.1 (1.1) 3.48 (1.35) 5.22 (0.63)

y HA 115.7 (18.6) 107.4 (36.3) 40 (38) 93.8 (45.7) 43.9 (25.7)

% 28 19 5 33 1

% 4 2 1 11 0

r 4 2.11 (0.21) 2.17 (0.27) 4.99 (0.64) 2.04 (0.28) 2.18 (0.58)

y 4 153.0 (12.9) 154.2 (12.5) 159.1 (10.7) 159.5 (11.0) 155.0 (13.4)

r 5 2.98 (0.58) 2.77 (0.67) - - 3.13 (0.96)

y 5 119.1 (16.8) 130.2 (20.2) - - 129.4 (31.3)

r 6 2.08 (0.22) 2.07 (0.19) - - 2.31 (0.79)

y 6 153.4 (10.9) 150.1 (11.3) - - 149.4 (16.0)

This table lists key structural indexes fluctuations for theG5mutants, alongwith the controlmutantU7C in
reactant states. Data analysis was performed over the last 65 ns of each simulation with a 10 ps sampling
frequency. Distance and angles ( Fig. 2.10 ) are in ˚ and degrees, respectively. SDs are listed in parentheses.
Boldface font is used to highlight key quantities that are significantly altered with respect to the WT sim-
ulation uponmutation and that are discussed in the text. F is the in-line fitness index. 150 r HB and y HB are the
hydrogen bond length and angle for the general base step; defined by G12:N1 d C17:HO2 0 d C17:O2 0 .
r HA and y HA are the hydrogen bond length and angle for the general acid step; defined by C1.1:O5 0 d G8:
HO2 0 d G8:O2 0 . The hydrogenbond contact percentage for the above entries, defined as the percentage of
the snapshots in which r 3.0 ˚ and y 120 . The hydrogen bond contact percentage for the above
entries, defined as the percentage of the snapshots in which r 2.5 ˚ and y 150 .

5.1.2 Single mutations at the C3 and G8 positions
The C3 and G8 positions form a Watson-Crick base pair in the eHHR
structure 73,74 and they are important in stabilizing the active site structure
and positioning the 2 0 OH of G8 for acid catalysis. Here, we consider a series
of single mutations (C3U, G8A, G8I, and G8D) for which representative
hydrogen bond patterns are shown in Fig. 2.11 .