Mechanical design of the first proximal Ig domain of human cardiac titin revealed by single molecule force spectroscopy.
The elastic I-band part of muscle protein titin contains two tandem immunoglobulin (Ig) domain regions of distinct mechanical properties. Until recently, the only known structure was that of the I27 module of the distal region, whose mechanical properties have been reported in detail. Recently, the structure of the first proximal domain, I1, has been resolved at 2.1A. In addition to the characteristic beta-sandwich structure of all titin Ig domains, the crystal structure of I1 showed an internal disulfide bridge that was proposed to modulate its mechanical extensibility in vivo. Here, we use single molecule force spectroscopy and protein engineering to examine the mechanical architecture of this domain. In contrast to the predictions made from the X-ray crystal structure, we find that the formation of a disulfide bridge in I1 is a relatively rare event in solution, even under oxidative conditions. Furthermore, our studies of the mechanical stability of I1 modules engineered with point mutations reveal significant differences between the mechanical unfolding of the I1 and I27 modules. Our study illustrates the varying mechanical architectures of the titin Ig modules.
Department of Biological Sciences, Columbia University, New York, NY 10027, USA.
Hongbin
H
Li
Julio M
JM
Fernandez
D000595:Amino Acid Sequence; D064211:Connectin; D004220:Disulfides / Q000737:chemistry; D006801:Humans; D007136:Immunoglobulins / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry* / Q000276:immunology; D009206:Myocardium / Q000737:chemistry*; D011489:Protein Denaturation; D015202:Protein Engineering; D011494:Protein Kinases / Q000737:chemistry* / Q000276:immunology; D017434:Protein Structure, Tertiary; D013057:Spectrum Analysis / Q000379:methods*
D064211:Connectin; D004220:Disulfides; D007136:Immunoglobulins; D009124:Muscle Proteins; C577285:TTN protein, human; D011494:Protein Kinases