title: An experimentally based computer search identifies unstructured membrane-binding sites in proteins: application to class I myosins, PAKS, and CARMIL.
abstract: Programs exist for searching protein sequences for potential membrane-penetrating segments (hydrophobic regions) and for lipid-binding sites with highly defined tertiary structures, such as PH, FERM, C2, ENTH, and other domains. However, a rapidly growing number of membrane-associated proteins (including cytoskeletal proteins, kinases, GTP-binding proteins, and their effectors) bind lipids through less structured regions. Here, we describe the development and testing of a simple computer search program that identifies unstructured potential membrane-binding sites. Initially, we found that both basic and hydrophobic amino acids, irrespective of sequence, contribute to the binding to acidic phospholipid vesicles of synthetic peptides that correspond to the putative membrane-binding domains of Acanthamoeba class I myosins. Based on these results, we modified a hydrophobicity scale giving Arg- and Lys-positive, rather than negative, values. Using this basic and hydrophobic scale with a standard search algorithm, we successfully identified previously determined unstructured membrane-binding sites in all 16 proteins tested. Importantly, basic and hydrophobic searches identified previously unknown potential membrane-binding sites in class I myosins, PAKs and CARMIL (capping protein, Arp2/3, myosin I linker; a membrane-associated cytoskeletal scaffold protein), and synthetic peptides and protein domains containing these newly identified sites bound to acidic phospholipids in vitro.
authors.affiliation: Laboratory of Cell Biology, NHLBI, National Institutes of Health, Bethesda, Maryland 20892, USA. brzeskah@mail.nih.gov
authors.forename: Hanna
authors.initials: H
authors.lastname: Brzeska
authors.forename: Jake
authors.initials: J
authors.lastname: Guag
authors.forename: Kirsten
authors.initials: K
authors.lastname: Remmert
authors.forename: Susan
authors.initials: S
authors.lastname: Chacko
authors.forename: Edward D
authors.initials: ED
authors.lastname: Korn
mesh_terms: D000048:Acanthamoeba / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000465:Algorithms; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D051379:Mice; D009218:Myosins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020539:Sequence Analysis, Protein
chemical_list: D002352:Carrier Proteins; D010455:Peptides; D015800:Protozoan Proteins; D009218:Myosins