title: An S-alkylating reagent with positive charges as an efficient solubilizer of denatured disulfide-containing proteins.
abstract: A novel S-alkylating reagent, N-(3-bromopropyl)-N,N,N',N',N'-pentamethyl-1,3-propanedi(ammonium bromide) (TAP2-Br) which carries two positive charges in the molecule, was prepared to increase the solubility or to decrease the hydrophobicity of cysteine-containing denatured proteins (or peptides). S-Alkylation with TAP2-Br introduces two positive charges per cysteine residue, which will effectively shift the net charge of a protein in the positive direction. Disulfide-containing proteins, such as hen egg-white lysozyme, RNase A, BSA, and soybean trypsin inhibitor (Kunitz type), were reduced and S-alkylated with TAP2-Br to evaluate the potential of this reagent compared with other S-alkylating reagents such as monoiodoacetic acid, bromosuccinic acid and (3-bromopropyl)trimethylammonium bromide. The solubilities of these denatured proteins in the pH range of 2-10 indicated that S-alkylation with TAP2-Br effectively solubilized not only basic proteins (lysozyme and RNase) but also an acidic protein containing a fairly large number of cysteine residues (BSA). Moreover, the retentions of cysteine-containing tryptic peptides derived from lysozyme on reversed-phase HPLC were greatly reduced by S-alkylation with TAP2-Br. These results indicate that TAP2-Br is very useful to increase the solubility of some cysteine-containing denatured proteins and to decrease the hydrophobicity of peptides containing cysteine residue(s).
authors.affiliation: Department of Bioengineering Science, Faculty of Engineering, Okayama University.
authors.forename: H
authors.initials: H
authors.lastname: Yamada
authors.forename: M
authors.initials: M
authors.lastname: Seno
authors.forename: A
authors.initials: A
authors.lastname: Kobayashi
authors.forename: T
authors.initials: T
authors.lastname: Moriyama
authors.forename: M
authors.initials: M
authors.lastname: Kosaka
authors.forename: Y
authors.initials: Y
authors.lastname: Ito
authors.forename: T
authors.initials: T
authors.lastname: Imoto
mesh_terms: D000477:Alkylating Agents / Q000138:chemical synthesis* / Q000737:chemistry*; D000478:Alkylation; D002412:Cations; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry*; D007700:Kinetics; D010084:Oxidation-Reduction; D010449:Peptide Mapping / Q000379:methods; D011489:Protein Denaturation; D011506:Proteins / Q000737:chemistry*; D000644:Quaternary Ammonium Compounds / Q000138:chemical synthesis* / Q000737:chemistry*; D012995:Solubility
chemical_list: D000477:Alkylating Agents; D002412:Cations; D004220:Disulfides; D011506:Proteins; D000644:Quaternary Ammonium Compounds; C092444:N-(3-bromopropyl)-N,N,N'N',N'-pentamethyl-1,3-propanediammonium; D003545:Cysteine