Conformational Changes in Human Arf1 on Nucleotide Exchange and Deletion of Membrane-binding Elements

Ronald D. Seidel, Juan Carlos Amor, Richard A. Kahn, James H. Prestegard
2004 Journal of Biological Chemistry  
Conformational changes associated with nucleotide exchange or truncation of the N-terminal ␣-helix of human Arf1 have been investigated by using forms of easily acquired NMR data, including residual dipolar couplings and amide proton exchange rates. ADP-ribosylation factors (Arfs) are 21-kDa GTPases that regulate aspects of membrane traffic in all eukaryotic cells. An essential component of the biological actions of Arfs is their ability to reversibly bind to membranes, a process that involves
more » ... cess that involves exposure of the myristoylated N-terminal amphipathic ␣-helix upon activation and GTP binding. Deletion of this helix results in a protein, termed ⌬17Arf1, that has a reduced affinity for GDP and the ability to bind GTP in the absence of lipids or detergents. Previous studies, comparing crystal structures for Arf1⅐GDP and ⌬17Arf1⅐GTP, identified several regions of structural variation and suggested that these be associated with nucleotide exchange rather than removal of the N-terminal helix. However, separation of conformational changes because of nucleotide binding and N-terminal truncation cannot be addressed in comparing these structures, because both the bound nucleotide and the N terminus differ. Resolving the two effects is important as any structural changes involving the N terminus may represent membrane-mediated conformational adjustments that precede GTP binding. Results from NMR experiments presented here on Arf1⅐GDP and ⌬17Arf1⅐GDP in solution reveal substantial structural differences that can only be associated with N-terminal truncation.
doi:10.1074/jbc.m402109200 pmid:15308674 fatcat:l7iqj5iyvrbpxmlztxo7q5qle4