title: Affinity, avidity, and kinetics of target sequence binding to LC8 dynein light chain isoforms.
abstract: LC8 dynein light chain (DYNLL) is a highly conserved eukaryotic hub protein with dozens of binding partners and various functions beyond being a subunit of dynein and myosin Va motor proteins. Here, we compared the kinetic and thermodynamic parameters of binding of both mammalian isoforms, DYNLL1 and DYNLL2, to two putative consensus binding motifs (KXTQTX and XG(I/V)QVD) and report only subtle differences. Peptides containing either of the above motifs bind to DYNLL2 with micromolar affinity, whereas a myosin Va peptide (lacking the conserved Gln) and the noncanonical Pak1 peptide bind with K(d) values of 9 and 40 μM, respectively. Binding of the KXTQTX motif is enthalpy-driven, although that of all other peptides is both enthalpy- and entropy-driven. Moreover, the KXTQTX motif shows strikingly slower off-rate constant than the other motifs. As most DYNLL partners are homodimeric, we also assessed the binding of bivalent ligands to DYNLL2. Compared with monovalent ligands, a significant avidity effect was found as follows: K(d) values of 37 and 3.5 nM for a dimeric myosin Va fragment and a Leu zipper dimerized KXTQTX motif, respectively. Ligand binding kinetics of DYNLL can best be described by a conformational selection model consisting of a slow isomerization and a rapid binding step. We also studied the binding of the phosphomimetic S88E mutant of DYNLL2 to the dimeric myosin Va fragment, and we found a significantly lower apparent K(d) value (3 μM). We conclude that the thermodynamic and kinetic fine-tuning of binding of various ligands to DYNLL could have physiological relevance in its interaction network.
authors.affiliation: Department of Biochemistry, Eötvös Loránd University, H-1117 Budapest, Hungary.
authors.forename: László
authors.initials: L
authors.lastname: Radnai
authors.forename: Péter
authors.initials: P
authors.lastname: Rapali
authors.forename: Zsuzsa
authors.initials: Z
authors.lastname: Hódi
authors.forename: Dániel
authors.initials: D
authors.lastname: Süveges
authors.forename: Tamás
authors.initials: T
authors.lastname: Molnár
authors.forename: Bence
authors.initials: B
authors.lastname: Kiss
authors.forename: Bálint
authors.initials: B
authors.lastname: Bécsi
authors.forename: Ferenc
authors.initials: F
authors.lastname: Erdödi
authors.forename: László
authors.initials: L
authors.lastname: Buday
authors.forename: József
authors.initials: J
authors.lastname: Kardos
authors.forename: Mihály
authors.initials: M
authors.lastname: Kovács
authors.forename: László
authors.initials: L
authors.lastname: Nyitray
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D056445:Cytoplasmic Dyneins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D009154:Mutation; D018995:Myosin Heavy Chains / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D024701:Myosin Type V / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization
chemical_list: D007527:Isoenzymes; D008024:Ligands; D010455:Peptides; C077511:MYO5A protein, human; C427758:DYNLL1 protein, human; D024701:Myosin Type V; D018995:Myosin Heavy Chains; D056445:Cytoplasmic Dyneins; C501531:DYNLL2 protein, human