title: Stability and folding rates of domains spanning the large A-band super-repeat of titin.
abstract: Titin is a very large (>3 MDa) protein found in striated muscle where it is believed to participate in myogenesis and passive tension. A prominent feature in the A-band portion of titin is the presence of an 11-domain super-repeat of immunoglobulin superfamily and fibronectin-type-III-like domains. Seven overlapping constructs from human cardiac titin, each consisting of two or three domains and together spanning the entire 11-domain super-repeat, have been expressed in Escherichia coli. Fluorescence unfolding experiments and circular dichroism spectroscopy have been used to measure folding stabilities for each of the constructs and to assign unfolding rates for each super-repeat domain. Immunoglobulin superfamily domains were found to fold correctly only in the presence of their C-terminal fibronectin type II domain, suggesting close and possibly rigid association between these units. The domain stabilities, which range from 8.6 to 42 kJ mol(-1) under physiological conditions, correlate with previously reported mechanical forces required to unfold titin domains. Individual domains vary greatly in their rates of unfolding, with a range of unfolding rate constants between 2.6 x 10(-6) and 1.2 s(-1). This variation in folding behavior is likely to be an important determinant in ensuring independent folding of domains in multi-domain proteins such as titin.
authors.affiliation: Department of Biochemistry, University of Bristol, Bristol BS8 1TD, United Kingdom.
authors.forename: J G
authors.initials: JG
authors.lastname: Head
authors.forename: A
authors.initials: A
authors.lastname: Houmeida
authors.forename: P J
authors.initials: PJ
authors.lastname: Knight
authors.forename: A R
authors.initials: AR
authors.lastname: Clarke
authors.forename: J
authors.initials: J
authors.lastname: Trinick
authors.forename: R L
authors.initials: RL
authors.lastname: Brady
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D002942:Circular Dichroism; D064211:Connectin; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D011489:Protein Denaturation; D017510:Protein Folding; D011494:Protein Kinases / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics
chemical_list: D000911:Antibodies, Monoclonal; D064211:Connectin; D009124:Muscle Proteins; C577285:TTN protein, human; D011494:Protein Kinases