Projects
I am interested in the
controls on the degradation of organic carbon by heterotrophic microbes in
marine environments. My focus has
primarily been on the dynamics of polysaccharides, monosaccharides, and
microbial extracellular enzymes in the water column. My graduate work in the lab of Dr. Carol Arnosti
can be broken into three main projects:
Polysaccharide hydrolysis and
monomer uptake in the marine water column
High molecular weight
dissolved organic carbon (DOC) is, on average, more available to heterotrophic
microbes than low molecular weight DOC.
To access large molecules, microbes must first hydrolyze them outside of
the cell with extracellular enzymes.
The presence of appropriate extracellular enzymes is therefore a
prerequisite for the consumption of much of the most labile DOC in the ocean.
Using fluorescently-labeled
polysaccharides to probe the ability of microbial communities to access
structurally specific polysaccharides, and using radiolabeled glucose to
measure the ability of those communities to access the products of
polysaccharide hydrolysis by extracellular enzymes, it is possible to evaluate
the spectrum of polysaccharides available to the microbial community and the
extent to which polysaccharide hydrolysis is a rate-limiting step in the remineralization
of dissolved organic carbon.
Steen, A.
D., L. Hamdan, and C. Arnosti.
2008. Limnology and Oceanography
53(3):936-947
Microbial turnover of high molecular
weight dissolved organic matter: Where are the roadblocks?
Steen,
A.D., K. Ziervogel, S. Ghobrial, and C. Arnosti.
2009. Talk delivered at the
2009 Aquatic Sciences meeting,
Steen,
A.D., and C. Arnosti. 2009. Poster presented at the symposium on
Chemical Oceanography in a Changing World,
Active lifetimes of extracellular
enzymes in seawater
In order for production of
extracellular enzymes to represent a viable strategy for obtaining resources,
microbes must "profit" from their production: that is, the enzymes
must return more resources than are required to synthesize the enzymes. The lifetime of extracellular enzymes in
seawater is a critical factor determining this balance, but it is almost
completely unconstrained.
Measurements from the Arctic (
Degradation rates of extracellular
enzymes in Arctic seawater
Steen, A.
D. and C. Arnosti. Talk delivered at the 2008 Ocean
Sciences meeting,
Novel methods to study
polysaccharide dynamics in aquatic environments
Most measurements of
polysaccharide hydrolase activities in seawater are based on "fluorogenic
small substrate proxies", small molecules consisting of a fluorophore
covalently bound to a monomer such as glucose. While that bond is intact, the
fluorophore is dark, and when the bond is hydrolyzed, the bond fluoresces. Enzyme activity is measured as the
rate of increase of fluorescence as a function of time. This method has the serious drawback
that the structure of small substrate proxies is much simpler than the complex,
three-dimensional structures of polysaccharides, and that structure is critical
in determining the interactions between the substrate and corresponding
enzyme. Some polysaccharide
hydrolases are completely incapable of hydrolyzing the corresponding small
substrate proxy - for instance, commercially available pullulanase, an α-glucosidase, does not hydrolyze methylumbelliferyl-α-d-glucopyranoside. I have helped to develop two methods of
measuring polysaccharide hydrolysis in aquatic samples that use labeled
polysaccharides. These methods are
rapid, use specific polysaccharides, and in addition to determining hydrolysis
rates can yield information about other dynamics of polysaccharides, such as
sorption to mineral surfaces.
Steen,
A.D., P. Gururaj, J. Ma, N.V. Blough, and C. Arnosti.
2008. Analytical
Biochemistry 383:340-342.
Steen,
A.D., C. Arnsoti, L. Ness, and N.V. Blough.
2006. Marine Chemistry
101:266-276