Our long-term goal is to understand the detailed mechanism of cytochrome c oxidase (ccO), one of the
most important proton pumps that has been studied extensively. X-ray and mutagenesis studies have
revealed two channels that facilitate proton translocation within the protein and are believed to be important
in different stages of the catalytic cycle. However, the precise relationship between proton pumping and
the reactions at the redox site is not understood and different mechanisms have been proposed. For
example, the classical paradigm suggests that all four protons are pumped after the initial reduction of the
binuclear center and the binding of O2. This mechanism was recently challenged by Michel, who
proposed, based on the re-evaluation of available experimental data and the principle of electroneutrality,
that one proton is pumped before O2 binding. Also the proton acceptors in several proposed intermediates
are not known, and it is not clear whether protons associated with them are "chemical" (for O2 reduction)
or "physical" (for pumping). The precise role of the copper ion in the proton pumping process is also not
well established. Careful computational work on the energetics of proposed pathways and dynamics of
ccO will be of great value in complementing experimental investigations on these mechanistic issues.
One system of great interest is the Cu, Zn-superoxide dismutase (CZSOD), an enzyme that employs
Cu/Zn to convert superoxide to dioxygen and hydrogen peroxide and therefore is the major defense against
superoxide. Strikingly, one mutation (e.g. A4V) can transform this antioxidant into a toxic pro-oxidant
that causes amyotrophic lateral sclerosis (ALS). Recently, it was found that ALS-related mutations cause a
dramatic loss in the Zn binding affinity of CZSOD. The Zn-deficient CZSOD was proposed to be a
stronger oxidant, and can react with cellular reductants and generates superoxide, which may react with
NO to form a strong pro-oxidant that was sufficient to induce apoptosis in cultured neurons. In earlier
works, however, it was found that Zn-deficient Cu-apoSOD has a similar reduction rate as the wild type.
It was proposed that the major role of the zinc ion is to facilitate the product release via an internal
displacement reaction, forcing the superoxide into an axial position, which is suitable for dissociation. To
better understand the role of the zinc ion in CZSOD, computational work will be carried out to probe the
redox property of the copper site, pK of active site residues and energetics of relevant chemical steps for
both the wild type enzyme and mutants known to be involved in ALS. The results will yield a detailed
view on the function of CZSOD, and possibly some general features of metalloenzymes that employ more
than one metal ion in the active site.
An essential element in bioenergetics is proton pumping, in which protons are driven from one side of
the membrane to the other, coupled with light-induced excitation (e.g. bacteriorhodopsin) or redox coupled
processes involving transition metal ions (e.g. cytochrome c oxidase). The resulting concentration
gradient is employed by other energy consuming processes such as ATP synthesis, as in the ATP synthase
above. Therefore, proton pumps can be thought as the "power engines" of living systems. Due to their
importance in bioenergetics, proton pumps have attracted much attention both experimentally and
theoretically. Although concepts such as "proton wires" are well accepted and characterized in a number of
systems, more issues such as the coupling between electron transfer, chemistry at redox sites and proton
translocation are still under active research and debate. Computational studies beyond classical MD should
be able to provide useful insights into the working mechanisms of those complicated systems.
The role of transition metals in neuroscience have not been recognized until fair recently, mainly driven
by the explosive number of publications pointing to their importance in the regulation of radicals that are
associated with many neurological diseases. Therefore, it is of great value to understand the specific role
of those metal ions in radical chemistry, and how are their functions regulated by the sequence, structure
and dynamics of the biomolecules to which they bind. More perplexing is the role of redox-inactive ions
such as zinc and aluminum. Although they are not capable of generating radical species directly, they are
present in many proteins that are relevant to neurodegenerative disorders (e.g. metallothionein) and are
believed to participate in many reactions indirectly. Is their role mainly structural, or do they influence the
redox properties of other metals through electrostatic interaction? How are the binding properties of the
metal ions regulated, and what is the consequence of their binding on protein conformations? These are
questions that are ideal for computational studies.