Professor
Biochemistry and molecular biology of biotin and biotin-containing enzymes
Regulation of plant lipid metabolism
Dr. Nikolau's research is focused on three projects in
the area of plant biochemistry and molecular genetics.
Biotin and biotin-containing enzymes
Biotin is a water-soluble vitamin that is biosynthesized by
plants and some bacteria and fungi. Its biochemical function is as a
covalently-bound cofactor on a family of enzymes that catalyze reactions
in a variety of crucial metabolic processes. Examples of such enzymes
are acetyl-CoA carboxylase, methylcrotonyl-CoA carboxylase and geranoyl-CoA
carboxylase, which are required for lipogenesis, leucine metabolism
and isoprenoid metabolism, respectively. Until recently, little was
known about the structure, function and regulation of biotin biosynthesis
and biotin-containing enzymes of plants. In the last five years Dr.
Nikolau's laboratory, in collaboration with Dr. Eve S. Wurtele, has
made major advances in the isolation and characterization of the genes
coding for biotin-containing enzymes and the enzymes required for biotin
biosynthesis.
Plants contain two isozymes of acetyl-CoA carboxylase that are located
in separate subcellular compartments, plastids and the cytosol. The
former is required for de novo fatty acid biosynthesis, whereas the
latter is required for the biosynthesis of a variety of seconary phytochemicals.
In most plants these two isozymes have unique quaternary structures.
The plastidic enzyme is heteromeric, being composed of four distinct
subunits; Dr. Nikolau's laboratory has isolated the genes coding for
all four of these subunits. The cytosolic acetyl-CoA carboxylase is
homomeric, being a dimer of identical subunits; Dr. Nikolau's laboratory
has isolated the genes coding for this subunit.
Methylcrotonyl-CoA carboxylase is a mitochondrial biotin-containing
enzyme involved in leucine catabolism. The enzyme is composed of two
different types of subunits in a dodecameric quaternary structure. Dr.
Nikolau's laboratory has isolated the genes coding for both of these
subunits.
Biotin synthase is the enzyme that catalyzes the terminal reaction of
biotin biosynthesis. The characterization of this enzyme has proven
to be extremely difficult. However, Dr. Nikolau's laboratory has recently
isolated the gene coding for biotin synthase of plants. This achievement
should enable the further characterization of biotin synthase, and of
biotin biosynthesis in plants.
Research is currently focused on the biochemical and genetic regulation
of these enzymes and genes. The elucidation of how these enzymes are
regulated will have impact on comprehending how each metabolic process
is controlled.
Lipid metabolism
Research is focused on the understanding the biosynthesis of unusual
plant lipids; specifically, cuticular waxes. Cuticular waxes are the
surface lipids that act as a water-barrier for the ariel parts of plants.
These lipids are derivatives of very long-chain fatty acids that are
synthesized by the epidermal cells of the plant. Molecular genetic approaches
are being taken to isolate genes required for the normal biosynthesis
of the cuticular waxes. In maize, at least 15 genes have been defined
by mutations that affect the normal accumulation of cuticular waxes.
These are termed glossy. In collaboration with Dr. Patrick S. Schnable,
transposon-tagging has been used to isolate the genes glossy1 and glossy8.
InArabidopsis a similar set of mutants, called cer, define genes required
for cuticular wax biosynthesis in this plant. The CER2 locus of Arabidopsis
has been cloned via chromosome walking. Research is now focused on elucidating
the biochemical function of the proteins encoded by these isolated genes.
The long term goal is to fully elucidate the cuticular wax biosynthetic
pathway and its biochemical and genetic regulation.
Genome structure and meiotic recombination
Meiotic recombination is a major mechanisms by which genetic diversity
is generated in a genome. Such diversity is a prerequisite for selection,
by which, the evolutionary development of a genome occurs. Although
meiotic recombination is dependent upon the physical organization of
the genome, this interrelationship is undefined. Dr. Nikolau's laboratory,
in collaboration with Dr. Patrick S. Schnable, is undertaking research
to examine the relationship between genome structure and meiotic recombination.
Research is focusing on a genetically defined interval of the maize
genome located between the a1 and sh2 loci. These two loci are separated
by a genetic interval of about 0.1 cM. Dr. Nikolau and collaborators
have determined that this genetic interval is equivalent to 140-kb of
DNA, which has been cloned in an Yeast Artifical Chromosome (YAC). Research
is now focused on determining the physical features of the 140-kb interval
that separate the a1 and sh2 loci, and mapping the location of about
200 meiotic recombination events that have been generated within this
same interval. The long-term goal is to define mechanistically how sites
of recombination are selected.
