LINK BETWEEN DYNAMICS AND FUNCTION IN SINGLE AND MULTI-SUBUNIT ENZYMES
LINK BETWEEN DYNAMICS AND FUNCTION IN SINGLE AND MULTI-SUBUNIT ENZYMES
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Date
2010
Authors
Chen, Jie
Advisor
Thirumalai, Devarajan
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Abstract
Biopolymers, such as proteins and DNA, are polymers whose three-dimensional
conformations dene their biological functions. Current emphasis on structures has
greatly advanced our understanding of the functions of biopolymers. However, there
is a need to understand the deeper link between biopolymer dynamics and function,
because in water and under cellular conditions, everything that biopolymers do can
be understood in terms of "the jigglings and wigglings of atoms". These motions
arise from thermal noise in the solvent and due to intrinsic motion of the enzymes. In
biological systems, the motions are often highly regulated to ensure that cellular processes are executed over the required time scales. For enzymes, which are essentially
proteins that catalyze chemical reactions or generate mechanical work, conformational
fuctuations are coupled at various stages through interactions with ligands
during the catalytic cycle. We have studied two dierent enzymes, dihydrofolate
reductase (DHFR), which catalyzes reduction of dihydrofolate to tetrahydrofolate,
and RNA polymerase (RNAP from bacteria and Pol II from yeast), which is responsible for RNA synthesis using DNA as a template. In order to study the link between
dynamics and function we have developed new methods and extended a variety of
computational techniques. For DHFR, we use both evolutionary imprints (SCA)
and structure-based perturbation method (SPM) to extract a network of residues
that facilitate the transitions between two distinct conformational states (closed and
occluded states). The transition kinetics and pathways connecting the closed and
occluded states are described using Brownian dynamics (BD) simulation. We found
the sliding motion of Met20 loop across helix 2 is involved in the forward and reverse
transitions between the closed and occluded states. We also found that cross-linking
M16-G121 inhibits both the forward and the reverse transitions. In addition, we
showed the transition states of these transitions are broad and resemble high energy
states.
For RNAP, we focus on the conformational changes of RNAP and DNA in
promoter melting process. Using BD, we show that DNA conformation changes
in promoter melting occur in three steps. We also show that internal dynamics
of RNAP is relevant to facilitate the bending of DNA. For Pol II, the structural
transitions between two initiation conformational states and between initiation state
and elongation state are studied using SPM and BD. We determine the structural
units that regulate structural transitions and describe the transition kinetics. The
combination of three dierent methods, SCA, SPM and BD, provide results that
are in accord with many experiments. Moreover, our description of the detailed
structural transitions in these enzymes lead to new insights and testable predictions
in these extraordinarily important enzyme functions.