Development of a Molecular Probe for Investigating Cell- Surface Enzyme Activity in Seawater

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Nitrogen is one of the nutrients essential for phytoplankton growth. The rate of nitrogen recycling can determine the rate of primary production. One of the major pathways of nitrogen cycling in the sea is the uptake of N by phytoplankton for incorporation into the amino acids and nucleic acids required for life. Traditionally, marine scientists have thought that phytoplankton obtain the N they require from inorganic nitrogen compounds. However, dissolved organic nitrogen compounds make up a larger reservoir of N in seawater than do inorganic nitrogen compounds. Previously, field studies have indicated that amino acids and other organic nitrogen compounds are not taken up to any significant extent by phytoplankton. It has been recently suggested that some phytoplankton can oxidize amines and amino acids using a cell-surface enzyme. Oxidation of amino acids produces hydrogen peroxide, an x-keto acid, and ammonium. The ammononium is then transported across the phytoplankton cell membrane for use as a N source. The x- keto acid is released to the surrounding seawater where it can be taken up by bacteria. If the ability to oxidize amino acids by mechanism is widespread among phytoplankton, it has profound implications for our understanding of the cycling of amino acids in seawater and the competition between phytoplankton and bacteria for these compounds. This project will develop a series of fluorescent molecular probes to simulate amino acids and amines in seawater. Loss of the probes (by algal oxidation) from seawater can be monitored quickly and sensitively by high performance liquid chromatography with fluorescent detection. Using these synthesized probes, the importance of phytoplankton oxidation of amines and amino acids by this novel mechanism will be tested in the field.