title: Probing actin and liposome interaction of talin and talin-vinculin complexes: a kinetic, thermodynamic and lipid labeling study.
abstract: Talin purified from human platelets and chicken gizzard smooth muscle is an actin and lipid binding protein. Here, we have investigated the effect of vinculin on (a) talin-nucleated actin polymerization and (b) insertion of talin into lipid bilayers. Calorimetric data show ternary complex formation between talin, vinculin, and actin. Actin-talin, actin-vinculin and actin-(talin-vinculin) binding and rate constants as well as actin polymerization rates for all three protein species have been determined by steady state titration, stopped-flow, and fluorescence assay. In contrast to an increase of the polymerization rate by a factor of less than 2 for actin-talin and actin-(talin-vinculin) when lowering the temperature, we measured a decrease in rates for actin alone and actin-vinculin. The overall equilibrium constants (Keq) in the van't Hoff plot proved linear and were of one-step reactions. Thermodynamic data exhibited signs of van der Waal's binding forces. Using the photoactivatable lipid analogue [3H]PTPC/11, which selectively labels membrane-embedded hydrophobic domains of proteins, we also show that talin partially inserts into the hydrophobic bilayer of liposomes. This insertion occurs in a similar manner irrespective of preincubation with vinculin.
authors.affiliation: Biophysics Department, Technical University of Munich, Garching, FRG.
authors.forename: W H
authors.initials: WH
authors.lastname: Goldmann
authors.forename: V
authors.initials: V
authors.lastname: Niggli
authors.forename: S
authors.initials: S
authors.lastname: Kaufmann
authors.forename: G
authors.initials: G
authors.lastname: Isenberg
mesh_terms: D000199:Actins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D001792:Blood Platelets / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008081:Liposomes / Q000737:chemistry*; D046911:Macromolecular Substances; D010718:Phosphatidylserines / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D016608:Talin / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D013816:Thermodynamics; D016596:Vinculin / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D000199:Actins; D008081:Liposomes; D046911:Macromolecular Substances; D010718:Phosphatidylserines; D016608:Talin; D016596:Vinculin