title: Requirements for the catalytic cycle of the N-ethylmaleimide-Sensitive Factor (NSF).
abstract: The N-ethylmaleimide-Sensitive Factor (NSF) was one of the initial members of the ATPases Associated with various cellular Activities Plus (AAA(+)) family. In this review, we discuss what is known about the mechanism of NSF action and how that relates to the mechanisms of other AAA(+) proteins. Like other family members, NSF binds to a protein complex (i.e., SNAP-SNARE complex) and utilizes ATP hydrolysis to affect the conformations of that complex. SNAP-SNARE complex disassembly is essential for SNARE recycling and sustained membrane trafficking. NSF is a homo-hexamer; each protomer is composed of an N-terminal domain, NSF-N, and two adjacent AAA-domains, NSF-D1 and NSF-D2. Mutagenesis analysis has established specific roles for many of the structural elements of NSF-D1, the catalytic ATPase domain, and NSF-N, the SNAP-SNARE binding domain. Hydrodynamic analysis of NSF, labeled with (Ni(2+)-NTA)(2)-Cy3, detected conformational differences in NSF, in which the ATP-bound conformation appears more compact than the ADP-bound form. This indicates that NSF undergoes significant conformational changes as it progresses through its ATP-hydrolysis cycle. Incorporating these data, we propose a sequential mechanism by which NSF uses NSF-N and NSF-D1 to disassemble SNAP-SNARE complexes. We also illustrate how analytical centrifugation might be used to study other AAA(+) proteins.
authors.affiliation: Department of Cellular and Molecular Biochemistry, University of Kentucky Medical Center, Lexington, KY 40536-0509, USA. chz024@ucsd.edu
authors.forename: Chunxia
authors.initials: C
authors.lastname: Zhao
authors.forename: Everett C
authors.initials: EC
authors.lastname: Smith
authors.forename: Sidney W
authors.initials: SW
authors.lastname: Whiteheart
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry; D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D000818:Animals; D006801:Humans; D006868:Hydrolysis; D008561:Membrane Fusion; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D052066:N-Ethylmaleimide-Sensitive Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D050600:SNARE Proteins / Q000378:metabolism; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins / Q000378:metabolism
chemical_list: D046912:Multiprotein Complexes; D050600:SNARE Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D000255:Adenosine Triphosphate; D052066:N-Ethylmaleimide-Sensitive Proteins