Protein Dynamics and Enzyme Functioning

Gekko, K. Compressibility gives new insight into protein dynamics and enzyme function, Biochim. Biophys. Acta. 1595 (2002) 382-386. 

FRET is an extremely useful phenomenon when it comes to the analysis of molecular conformations and interactions. F or the analysis of interactions, in which two separate molecules are labeled with an appropriate pair of fluorophores, an interaction can be shown by observing FRET. Further, FRET can be used as a type of spectroscopic ruler to measure the closeness of interactions. Proteins, lipids, enzymes, DNA, and RNA can all be labeled and interactions documented. This general method can be applied not only to questions of cellular function like kinase dynamics [3] but also to disease pathways, for example, the APP-PS1 interaction that is important in Alzheimer s disease (AD) [4], Alternatively, two parts of a molecule of interest can be labeled with a donor and acceptor fluorophore. Using this technique, changes in protein conformation and differences between isoforms of proteins can be measured, as well as protein cleavage. 

TTie diversity in the type of Fe-S clusters associated with these enzymes, catalyzing apparently simple hydration and/or dehydration reactions, is striking. Taken together, these results suggest that some of the Fe-S clusters that have been assigned redox roles in various enzymes may actually be functioning as catalytic groups. Clearly the field of Fe-S proteins, which a decade ago seemed to be well understood, has developed into a dynamic and fertile area for future research endeavors. 

In addition to structural roles, it is becoming increasingly apparent that a major function of IFs is to regulate the activities of cell metabolism, by serving as scaffolding for the dynamic regulation of associated proteins such as enzymes, signaling adapter molecules, stress proteins, cell death receptors, and even the endocytic machinery. 

Another important area of dynamic studies in biological samples is the effect of hydration upon molecular mobility in proteins and carbohydrates. The reason for these studies is primarily that protein dynamics, in particular, are crucial to their function, and so examining factors, such as the degree of hydration, that affect their dynamics is very important. However, it is obviously near-impossible to study dynamics in aqueous solution as a function of degree of hydration, and, since most proteins are not soluble in nonaqueous solvents, solid-state studies must be used. The motions at three methionine (Met) residues in Streptomyces subtilisin inhibitor (SSI) were studied with 2H NMR using a sample in which the Met residues at two crucial enzyme recognition sites (PI and P4) were specifically deuterated, along with one in the hydrophobic core.114 The motions of the Met side-chains were then examined... 

Biomolecular recognition is mediated by water motions, and the dynamics of associated water directly determine local structural fluctuation of interacting partners [4, 9, 91]. The time scales of these interactions reflect their flexibility and adaptability. For water at protein surfaces, the studies of melittin and other proteins [45, 46] show water motions on tens of picoseconds. For trapped water in protein crevices or cavities, the dynamics becomes much slower and could extend to nanoseconds [40, 71, 92], These rigid water molecules are often hydrogen bonded to interior residues and become part of the structural integrity of many enzymes [92]. Here, we study local water motions in various environments, from a buried crevice to an exposed surface using site-selected tryptophan but with different protein conformations, to understand the correlation between hydration dynamics and conformational transitions and then relate them to biological function. 

In recent years, increasing attention has been focused on proteins derived from extreme thermophylic bacteria (Daniel and Cowan, 2000 Vetriani et al., 1998 Jaenicke, 1996 1998, 2000 Adams and Kelly, 2001 and references therein). The increasing use of these proteins in biotechnology has given new impetus to studies focused on their structure and stability. At the same time, thermostable proteins prove challenging as the ideal candidates for investigating the relationships between the structure and intramolecular dynamics of the enzyme on the one hand, and their function and stability on the other. 

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