title: Analysis of the solution conformations of T4 lysozyme by paramagnetic NMR spectroscopy.
abstract: A large number of crystal structures of bacteriophage T4 lysozyme (T4-L) have shown that it contains two subdomains, which can arrange in a compact conformation (closed state) or, in mutants of T4-L, more extended structures (open state). In solution, wild-type T4-L displays only a single set of nuclear magnetic resonance (NMR) signals, masking any conformational heterogeneity. To probe the conformational space of T4-L, we generated a site-specific lanthanide binding site by attaching 4-mercaptomethyl dipicolinic acid via a disulfide bond to Cys44 in the triple-mutant C54T/C97A/S44C of T4-L and measured pseudocontact shifts (PCS) and magnetically induced residual dipolar couplings (RDC). The data indicate that, in solution and in the absence of substrate, the structure of T4-L is on average more open than suggested by the closed conformation of the crystal structure of wild-type T4-L. A slightly improved fit was obtained by assuming a population-weighted two-state model involving an even more open conformation and the closed state, but paramagnetic relaxation enhancements measured with Gd(3+) argue against such a conformational equilibrium. The fit could not be improved by including a third conformation picked from the hundreds of crystal structures available for T4-L mutants.
authors.affiliation: State Key Laboratory of Elemento-organic Chemistry, The Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin 300071, China. xunchengsu@nankai.edu.cn.
authors.forename: Jia-Liang
authors.initials: JL
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Elemento-organic Chemistry, The Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin 300071, China. xunchengsu@nankai.edu.cn.
authors.forename: Yin
authors.initials: Y
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Elemento-organic Chemistry, The Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin 300071, China. xunchengsu@nankai.edu.cn.
authors.forename: Lin-Lin
authors.initials: LL
authors.lastname: Zhang
authors.affiliation: Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia. t.huber@anu.edu.au gottfried.otting@anu.edu.au.
authors.forename: Haobo
authors.initials: H
authors.lastname: Liang
authors.affiliation: Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia. t.huber@anu.edu.au gottfried.otting@anu.edu.au.
authors.forename: Thomas
authors.initials: T
authors.lastname: Huber
authors.affiliation: State Key Laboratory of Elemento-organic Chemistry, The Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin 300071, China. xunchengsu@nankai.edu.cn and Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia. t.huber@anu.edu.au gottfried.otting@anu.edu.au.
authors.forename: Xun-Cheng
authors.initials: XC
authors.lastname: Su
authors.affiliation: Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia. t.huber@anu.edu.au gottfried.otting@anu.edu.au.
authors.forename: Gottfried
authors.initials: G
authors.lastname: Otting
mesh_terms: D017122:Bacteriophage T4 / Q000737:chemistry* / Q000201:enzymology*; D018360:Crystallography, X-Ray; D028581:Lanthanoid Series Elements / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D009113:Muramidase / Q000737:chemistry*; D011487:Protein Conformation; D012996:Solutions / Q000737:chemistry
chemical_list: D028581:Lanthanoid Series Elements; D012996:Solutions; D009113:Muramidase