Ma K, Kan L, Wang K
Laboratory of Physical Biology, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Biochemistry. 2001 Mar 27;40(12):3427-38. doi: 10.1021/bi0022792.
Titin is a family of giant elastic proteins that constitute an elastic sarcomere matrix in striated muscle. In the I-band region of the sarcomere, where titin extends and develops passive force upon stretch, titin is composed of tandem repeats of approximately 100 residue immunoglobin domains and approximately 28-residue PEVK modules. We have performed 2D NMR and circular dichroism (CD) studies of the conformations of one representative 28-mer PEVK module from human fetal titin (PEPPKEVVPEKKAPVAPPKKPEVPPVKV). NMR data of synthetic peptides of this module as well as three constituent peptides of 9 to 12 residues in aqueous solutions reveal distinguishing features for left-handed three-residue per turn PPII helices: the lack of NOE NN(i, i+1), very large NOE alphaN(i, i+1)/NN(i, i+1), no medium range NOE alphaN(i, i+2), and dihedral angles phi and psi values of -78 and 146, respectively. Structural determinations indicate the presence of three short stretches of PPII helices of 4, 5, and 6 residues that are interposed with an unordered, and presumably flexible, spacer region to give one "polyproline II helix-coil" or "PhC" motif for roughly every 10 residues. These peptides also display the characteristic PPII CD spectra: positive peak or negative shoulder band at 223 nm, negative CD band near 200 nm, and biphasic thermal titration curves that reflect varied stability of these PPII helices. We propose that this PhC motif is a fundamental feature and that the number, length, stability, and distribution of PPII is important in the understanding of the elasticity and protein interactions of the PEVK region of titin.
肌联蛋白是一类巨大的弹性蛋白家族，在横纹肌中构成弹性肌节基质。在肌节的I带区域，肌联蛋白在此伸展并在拉伸时产生被动力，它由大约100个残基的免疫球蛋白结构域和大约28个残基的PEVK模块串联重复组成。我们对来自人胎儿肌联蛋白的一个代表性28聚体PEVK模块（PEPPKEVVPEKKAPVAPPKKPEVPPVKV）的构象进行了二维核磁共振（NMR）和圆二色性（CD）研究。该模块以及三种由9至12个残基组成的合成肽在水溶液中的NMR数据揭示了左手三残基每圈PPII螺旋的显著特征：缺乏NOE NN(i, i + 1)、非常大的NOE alphaN(i, i + 1)/NN(i, i + 1)、没有中等范围的NOE alphaN(i, i + 2)以及二面角phi和psi值分别为-78和146。结构测定表明存在三段短的PPII螺旋，分别为4、5和6个残基，它们被一个无序且可能灵活的间隔区隔开，大约每10个残基形成一个“多聚脯氨酸II螺旋 - 卷曲”或“PhC”基序。这些肽还显示出特征性的PPII CD光谱：在223 nm处有正峰或负肩带、在200 nm附近有负CD带以及反映这些PPII螺旋不同稳定性的双相热滴定曲线。我们提出这个PhC基序是一个基本特征，并且PPII的数量、长度、稳定性和分布对于理解肌联蛋白PEVK区域的弹性和蛋白质相互作用很重要。
