Structure and tropomyosin binding properties of the N-terminal capping domain of tropomodulin 1.
Two families of actin regulatory proteins are the tropomodulins and tropomyosins. Tropomodulin binds to tropomyosin (TM) and to the pointed end of actin filaments and "caps" the pointed end (i.e., inhibits its polymerization and depolymerization). Tropomodulin 1 has two distinct actin-capping regions: a folded C-terminal domain (residues 160-359), which does not bind to TM, and a conserved, N-terminal region, within residues 1-92 that binds TM and requires TM for capping activity. NMR and circular dichroism were used to determine the structure of a peptide containing residues 1-92 of tropomodulin (Tmod1(1-92)) and to define its TM binding site. Tmod1(1-92) is mainly disordered with only one helical region, residues 24-35. This helix forms part of the TM binding domain, residues 1-35, which become more ordered upon binding a peptide containing the N-terminus of an alpha-TM. Mutation of L27 to E or G in the Tmod helix reduces TM affinity. Residues 49-92 are required for capping but do not bind TM. Of these, residues 67-75 have the sequence of an amphipathic helix, but are not helical. Residues 55-62 and 76-92 display negative 1H-15N heteronuclear Overhauser enhancements showing they are flexible. The conformational dynamics of these residues may be important for actin capping activity.
Department of Neuroscience and Cell Biology, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854-5635, USA. greenfie@rwja.umdnj.edu
Norma J
NJ
Greenfield
Alla S
AS
Kostyukova
Sarah E
SE
Hitchcock-DeGregori
D000199:Actins / Q000737:chemistry; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry*; D002645:Chickens; D002942:Circular Dichroism; D004247:DNA / Q000737:chemistry; D004586:Electrophoresis; D009682:Magnetic Resonance Spectroscopy; D008840:Microfilament Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry; D012680:Sensitivity and Specificity; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D051348:Tropomodulin; D014335:Tropomyosin / Q000737:chemistry*
D000199:Actins; D002352:Carrier Proteins; D008840:Microfilament Proteins; D010455:Peptides; D011506:Proteins; D051348:Tropomodulin; D014335:Tropomyosin; D004247:DNA