PARACOCCUS DENITRIFICANS IRON TRANSPORT APPARATUS

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This program will focus on the bioorganic chemistry of ferric parabactin processing by Paracoccus denitrificans, i.e., the microorganism's iron uptake apparatus. Special emphasis will be placed on the role of the ferric parabactin receptor. Our studies to date have revealed that 1) the kinetics of ferric parabactin promoted iron uptake are consistent with a "Taxi Cab" mechanism in which ferric parabactin is adsorbed to the surface of the microorganism, the iron liberated and the deferrated ligand released; 2) the microorganism has an 80kD outer membrane protein inducible by iron starvation that binds ferric L-parabactin with an apparent Kd of 0.7uM, a protein which we have substantially purified; 3) this protein does not release iron from ferric parabactin by itself; and 4) the photoaffinity label, ferric L-parabactin azide can be utilized to label the ferric parabactin receptor protein. With these findings in hand and with the techniques we have developed, we will address the following questions, questions which although not exclusively, are largely focused on the role on the ferric parabactin receptor. 1) How substrate specific is the ferric parabactin receptor? 2) Is deferrated ferric parabactin released as intact parabactin? 3) How is iron liberated from ferric parabactin after it has been bound to the receptor? 4) What structural features of the receptor protein are critical to ferric parabactin processing, i.e., what are the key structural components of the protein responsible for binding? The relevance of the ferric parabactin receptor protein is in part associated with the a recent interest in utilizing these surface expressed proteins in the development of vaccines for opportunistic bacteria. Unfortunately, little is known about the isolation and properties of these siderophore receptors, a situation which this program will alleviate at least in part.