Nathan Alder
 
    My research in the Theg lab focuses on elucidation of the energetic requirements of protein translocation via the DpH-dependent pathway in thylakoids.  Protein translocation pathways generally require the hydrolysis of nucleotide triphosphates to power the transport of proteins across membranes.  In contrast, the DpH-dependent pathway requires only a trans-membrane proton gradient (DpH) to fuel protein transport.  The ultimate aim of my project is to determine the cost in free energy of translocating proteins along the DpH-dependent pathway.
    Much of my work has involved in vitro protein import assays with isolated thylakoids in which the DpH across the thylakoid membrane is concurrently measured using a fluorescent dye.  This work has provided evidence of a energetic threshold below which protein transport does not occur, and above which protein transport is a linear function of energetic input.  Importantly, this threshold appears to be substrate-specific in that different values are obtained when the transport of different pre-protein substrates of this pathway are measured.
    The current focus of my investigation is twofold.  First, I am performing assays to test whether the trans-thylakoid proton gradient directly energizes protein transport, as may occur in a proton/protein antiporter mechanism.  Second, I am making proton flux measurements while performing in vitro protein import assays to gain a quantitative measure of the number of protons utilized from the gradient per protein translocated.  This measure, combined with the measured energetic threshold values for protein translocation, in theory provides the free energy change involved in protein translocation via the DpH-dependent pathway.  This would represent the first direct measure of the energetic requirement for trans-membrane protein translocation in eukaryotes.