title: Synthesis of l- and d-Ubiquitin by One-Pot Ligation and Metal-Free Desulfurization.
abstract: Native chemical ligation combined with desulfurization has become a powerful strategy for the chemical synthesis of proteins. Here we describe the use of a new thiol additive, methyl thioglycolate, to accomplish one-pot native chemical ligation and metal-free desulfurization for chemical protein synthesis. This one-pot strategy was used to prepare ubiquitin from two or three peptide segments. Circular dichroism spectroscopy and racemic protein X-ray crystallography confirmed the correct folding of ubiquitin. Our results demonstrate that proteins synthesized chemically by streamlined 9-fluorenylmethoxycarbonyl (Fmoc) solid-phase peptide synthesis coupled with a one-pot ligation-desulfurization strategy can supply useful molecules with sufficient purity for crystallographic studies.
authors.affiliation: School of Medical Engineering, Hefei University of Technology, Hefei, 230009, P. R. China.|State Key Laboratory of Medicinal Chemical Biology, NanKai University, 94 Weijin Road, Tianjin, 300071, P. R. China.|Department of Chemistry, School of Life Sciences, Tsinghua University, Beijing, 100084, P. R. China.
authors.forename: Yi-Chao
authors.initials: YC
authors.lastname: Huang
authors.affiliation: High Magnetic Field Laboratory, Chinese Academy of Sciences, Hefei, 230026, P. R. China.
authors.forename: Chen-Chen
authors.initials: CC
authors.lastname: Chen
authors.affiliation: Department of Chemistry, School of Life Sciences, Tsinghua University, Beijing, 100084, P. R. China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Gao
authors.affiliation: School of Medical Engineering, Hefei University of Technology, Hefei, 230009, P. R. China.|State Key Laboratory of Medicinal Chemical Biology, NanKai University, 94 Weijin Road, Tianjin, 300071, P. R. China.
authors.forename: Ye-Hai
authors.initials: YH
authors.lastname: Wang
authors.affiliation: School of Medical Engineering, Hefei University of Technology, Hefei, 230009, P. R. China.|State Key Laboratory of Medicinal Chemical Biology, NanKai University, 94 Weijin Road, Tianjin, 300071, P. R. China.
authors.forename: Hua
authors.initials: H
authors.lastname: Xiao
authors.affiliation: Department of Chemistry, School of Life Sciences, Tsinghua University, Beijing, 100084, P. R. China. wfeng@mail.tsinghua.edu.cn.
authors.forename: Feng
authors.initials: F
authors.lastname: Wang
authors.affiliation: High Magnetic Field Laboratory, Chinese Academy of Sciences, Hefei, 230026, P. R. China. cltian@ustc.edu.cn.
authors.forename: Chang-Lin
authors.initials: CL
authors.lastname: Tian
authors.affiliation: School of Medical Engineering, Hefei University of Technology, Hefei, 230009, P. R. China. lym2007@mail.ustc.edu.cn, ymli@hfut.edu.cn.|State Key Laboratory of Medicinal Chemical Biology, NanKai University, 94 Weijin Road, Tianjin, 300071, P. R. China. lym2007@mail.ustc.edu.cn, ymli@hfut.edu.cn.
authors.forename: Yi-Ming
authors.initials: YM
authors.lastname: Li
mesh_terms: D018360:Crystallography, X-Ray; D005449:Fluorenes / Q000737:chemistry; D008026:Ligation; D008968:Molecular Conformation; D010455:Peptides / Q000737:chemistry*; D060327:Solid-Phase Synthesis Techniques; D013438:Sulfhydryl Compounds / Q000737:chemistry; D025801:Ubiquitin / Q000138:chemical synthesis* / Q000737:chemistry
chemical_list: C496789:9-fluorenylmethoxycarbonyl; D005449:Fluorenes; D010455:Peptides; D013438:Sulfhydryl Compounds; D025801:Ubiquitin
keywords: X-ray diffraction; desulfurization; methyl thioglycolate; native chemical ligation; proteins