title: Molecular extensibility of mini-dystrophins and a dystrophin rod construct.
abstract: Muscular dystrophies arise with various mutations in dystrophin, implicating this protein in force transmission in normal muscle. With 24 three-helix, spectrin repeats interspersed with proline-rich hinges, dystrophin's large size is an impediment to gene therapy, prompting the construction of mini-dystrophins. Results thus far in dystrophic mice suggest that at least one hinge between repeats is necessary though not sufficient for palliative effect. One such mini-dystrophin is studied here in forced extension at the single molecule level. Delta2331 consists of repeats (R) and hinges (H) H1-R1-2 approximately H3 approximately R22-24-H4 linked by native (-) and non-native (approximately) sequence. This is compared to its core fragment R2 approximately H3 approximately R22 as well as an eight-repeat rod fragment middle (RFM: R8-15). We show by atomic force microscopy that all repeats extend and unfold at forces comparable to those that a few myosin molecules can generate. The hinge regions most often extend and transmit force while limiting tandem repeat unfolding. From 23-42 degrees C, the dystrophin constructs also appear less temperature-sensitive in unfolding compared to a well-studied betaI-spectrin construct. The results thus reveal new modes of dystrophin flexibility that may prove central to functions of both dystrophin and mini-dystrophins.
authors.affiliation: Pennsylvania Muscle Institute and Graduate Groups in Physics and Cell & Molecular Biology, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors.forename: Nishant
authors.initials: N
authors.lastname: Bhasin
authors.forename: Richard
authors.initials: R
authors.lastname: Law
authors.forename: George
authors.initials: G
authors.lastname: Liao
authors.forename: Daniel
authors.initials: D
authors.lastname: Safer
authors.forename: Jennifer
authors.initials: J
authors.lastname: Ellmer
authors.forename: Bohdana M
authors.initials: BM
authors.lastname: Discher
authors.forename: H Lee
authors.initials: HL
authors.lastname: Sweeney
authors.forename: Dennis E
authors.initials: DE
authors.lastname: Discher
mesh_terms: D000465:Algorithms; D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D016189:Dystrophin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011489:Protein Denaturation
chemical_list: D016189:Dystrophin