title: Dynamic motions of CD39 transmembrane domains regulate and are regulated by the enzymatic active site.
abstract: The two transmembrane domains flanking the active site of CD39 regulate its activity, but little is known about the structural and dynamic features underlying their importance. Here we use a disulfide crosslinking strategy to examine transmembrane helix interactions and dynamics and to correlate these features with activity and substrate binding. We find strong intrasubunit TM1-TM2 interactions, as well as TM1-TM1' and TM2-TM2' interactions between dimer subunits, near the extracellular side of the membrane but only weak interactions near the cytoplasmic end. The specific helix faces that constitute each interface are highly flexible, indicating a significant degree of rotational mobility within the packed structure. Analysis of activity after locking the helices in various orientations via disulfide bonds suggests that not only the arrangement but also the ability of the helices to move relative to each other is crucial for enzyme function. Helix mobility is in turn modulated by substrate binding. These results suggest that rather than playing a static structural role to support an optimal active site conformation, the transmembrane domains undergo dynamic motions that underlie their functional relationship with the active site.
authors.affiliation: Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts 02138, USA. agrinth@fas.harvard.edu
authors.forename: Alison
authors.initials: A
authors.lastname: Grinthal
authors.forename: Guido
authors.initials: G
authors.lastname: Guidotti
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001081:Apyrase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D019556:COS Cells; D003432:Cross-Linking Reagents / Q000378:metabolism; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D004789:Enzyme Activation / Q000235:genetics; D004795:Enzyme Stability; D016297:Mutagenesis, Site-Directed; D010618:Phenanthrolines / Q000378:metabolism; D017433:Protein Structure, Secondary / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051381:Rats; D013379:Substrate Specificity; D013816:Thermodynamics; D014162:Transfection
chemical_list: D015703:Antigens, CD; D003432:Cross-Linking Reagents; D010618:Phenanthrolines; D021122:Protein Subunits; D000251:Adenosine Triphosphatases; D001081:Apyrase; C067869:CD39 antigen; D003545:Cysteine; C025205:1,10-phenanthroline