Amino acid rotational spectra and internal friction in enzymes

1. Movements defining conformational changes are realized in changes of the psi and phi rotational angles of each amino acid. Using femtosecond resolution MD of proteins one can obtain the sub-picosecond dynamical characteristics of the individual amino acids. Defining rotational movements of the amino acids may be revealed by Fourier analysis of the rotational time series of the MD.

The spectral distributions of the psi rotations of the myosin can be grouped into 12 distinct groups. The central pie chart demonstrates the distribution of amino acids between these groups, the graphs at the sides show the spectral shape and variance of the individual groups. The same groups are present in calmodulin and trypsin rotational spectra, though with warying weight.

Experiments are carried out from mid-2011 in HZDR Dresden to test if these characteristic rotational patterns, may be induced by high intensity coherent irradiation of corresponding frequencies.

2. Upon activation of trypsinogen four peptide segments flanked by hinge glycine residues undergo conformational changes. To test whether the degree of conformational freedom of hinge regions affects the rate of activation, we introduced amino acid side chains of different characters at one of the hinges (position 193) and studied their effects on the rate constant of the conformational change. This structural rearrangement leading to activation was triggered by a pH-jump and monitored by intrinsic fluorescence change in the stopped-flow apparatus. We found that an increase in the size of the side chain at position 193 is associated with the decrease of the reaction rate constant. To analyze the thermodynamics of the reaction, temperature dependence of the reaction rate constants was examined in a wide temperature range (5-60 degrees C) using a novel temperature-jump/stopped-flow apparatus developed in our laboratory. Our data show that the mutations do not affect the activation energy (the exponential term) of the reaction, but they significantly alter the preexponential term of the Arrhenius equation. The effect of solvent viscosity on the rate constants of the conformational change during activation of the wild type enzyme and its R193G and R193A mutants was determined and evaluated on the basis of Kramers’ theory. Based on this we propose that the reaction rate of this conformational transition is regulated by the internal molecular friction, which can be specifically modulated by mutagenesis in the hinge region.