title: Identification of histidine tautomers in proteins by 2D 1H/13C(delta2) one-bond correlated NMR.
abstract: If the 13Cdelta2 chemical shift of neutral ("high pH") histidine is >122 ppm, primarily Ndelta1-H tautomer (2) is indicated; if it is <122 ppm, primarily Nepsilon2-H tautomer (1) is indicated. His resonances from the catalytic triad of active serine proteases, for example, are readily distinguished from those of denatured enzyme. The 13Cdelta2 chemical shifts increased by 6.2 ppm for the catalytic histidines in both alpha-lytic protease and subtilisin BPN' in raising the pH from that of imidazolium cation to that of tautomer 2. This tautomer identification method is easy to implement, requiring only bioincorporation of [U-13C] (or the more readily available [U-13C,15N])-histidine. Standard 1H/13C correlation HMQC or HSQC NMR pulse programs then yield the 13Cdelta2 chemical shifts with the benefit of high 1H sensitivity. Because of large one-bond spin-couplings (1JCH approximately 200 Hz), the method should extend to proteins having large 1H and 13C line widths, including very high molecular weights.
authors.affiliation: Department of Biochemistry, Tufts University School of Medicine, Boston, Massachusetts 02111, USA.
authors.forename: James L
authors.initials: JL
authors.lastname: Sudmeier
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: Bradshaw
authors.forename: Kristin E Coffman
authors.initials: KE
authors.lastname: Haddad
authors.forename: Regina M
authors.initials: RM
authors.lastname: Day
authors.forename: Craig J
authors.initials: CJ
authors.lastname: Thalhauser
authors.forename: Peter A
authors.initials: PA
authors.lastname: Bullock
authors.forename: William W
authors.initials: WW
authors.lastname: Bachovchin
mesh_terms: D000952:Antigens, Polyomavirus Transforming / Q000737:chemistry; D001665:Binding Sites; D002247:Carbon Isotopes; D006639:Histidine / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry*; D012697:Serine Endopeptidases / Q000737:chemistry; D013381:Subtilisins / Q000737:chemistry
chemical_list: D000952:Antigens, Polyomavirus Transforming; D002247:Carbon Isotopes; D011506:Proteins; D006639:Histidine; D012697:Serine Endopeptidases; D013381:Subtilisins; C021826:myxobacter alpha-lytic proteinase