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RESEARCH INTERESTS
Enzymology; DNA-Protein Interaction
Structure-function studies of gamma-glutamyl carboxylase
Gamma-glutamyl carboxylation and its role in growth & development of Drosophila
PUBLICATIONS
Ribosomal synthesis of proteins utilizes the available
standard amino acids. The functional repertoire of the
amino acids is greatly enhanced by post-translational
modification of some of the amino acids. For example,
reversible phosphorylation and dephosphorylation is used
in cellular signaling, acetylation and deacetylation in
remodeling chromatin structure, acylation for anchoring
proteins to membranes. Other modifications include
amidation of carboxy-terminus, hydroxylation of proline,
cis-trans isomerization of peptide bonds and L- to D-
isomerization of amino acids.
One such modification, gamma-carboxylation of
glutamate residues ( referred to as gamma-
carboxylation), is being investigated in the laboratory.
The reaction is carried out by the enzyme gamma-
glutamyl carboxylase and requires vitamin K as a
cofactor, hence the process is also referred to as
vitamin K-dependent carboxylation. In this process
glutamate (glu) residues are converted to gamma-
carboxy glutamate (gla). The modification was
discovered in proteins of blood coagulation cascade and
some bone proteins, and was thought to be mammalian
system-specific. Gla binds Ca++ and facilitates
the interaction of the protein with membranes. This
modification was subsequently identified in
neuropeptides synthesized in molluscs belonging to the
genus Conus suggesting a wider biological role of
gamma-carboxylation. The modifications are essential
for their interactions with cognate receptors on the cell
membrane. Glu residues in different sequence and
structural contexts are modified in the Conus
neuropeptides in contrast to the rather similar
sequences modified in the coagulation factors by the
mammalian enzyme. In addition the propeptide sequences
of these peptides which act as signals for carboxylation
are widely different for the Conus peptides, while they
are very similar for the mammalian substrate. Thus the
Conus enzyme offers a greater opportunity for studying
the structure-function relationship of the enzyme. We
have characterized a number of propeptide sequences
that can act as signals for gamma-carboxylation of
synthetic substrates by the Conus enzyme. Work is in
progress for expressing the enzyme in cells in culture.
This will enable us to synthesize both wild type and
enzymes carrying mutations for structure-function
studies.
We have recently demonstrated the presence of the
enzyme in Drosophila. The substrate requirements for
this enzyme are different from the Conus enzyme. We
are in the process of isolating mutations in the gene.
The genetic techniques available for studying Drosophila
can now be used to determine the wider role of gamma-
carboxylation in cellular processes. In addition,
availability of the mammalian, Conus, and Drosophila
enzymes will enable a comparative structure-function study of the enzyme.
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