Assignment of the side-chain sup 1 H and sup 13 C resonances of interleukin-1. beta. using double- and triple-resonance heteronuclear three-dimensional NMR spectroscopy

Journal Article · Tue Sep 04 00:00:00 EDT 1990 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00487a027· OSTI ID:5706499

Clore, G M; Bax, A; Driscoll, P C; Wingfield, P T; Gronenborn, A M ^[1]

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1. National Institute of Health, Bethesda, MD (USA)

The assignment of the aliphatic {sup 1}H and {sup 13}C resonances of IL-1{beta}, a protein of 153 residues and molecular mass 17.4 kDa, is presented by use of a number of novel three-dimensional (3D) heteronuclear NMR experiments which rely on large heteronuclear one-bond J couplings to transfer magnetization and establish through-bond connectivities. These 3D NMR experiments circumvent problems traditionally associated with the application of conventional 2D {sup 1}H-{sup 1}H correlation experiments to proteins of this size, in particular the extensive chemical shift overlap which precludes the interpretation of the spectra and the reduced sensitivity arising from {sup 1}H line widths that are often significantly larger than the {sup 1}H-{sup 1}H J couplings. The assignment proceeds in two stages. In the first step the {sup 13}C{alpha} chemical shifts are correlated with the NH and {sup 15}N chemical shifts by a 3D triple-resonance NH-{sup 15}N-{sup 13}C{alpha} (HNCA) correlation experiment which reveals both intraresidue NH(i)-{sup 15}N(i)-{sup 13}C{alpha}(i) and some weaker interresidue NH(i)-{sup 15}N(i)-C{alpha}(i-1) correlations, the former via intraresidue one-bond {sup 1}J{sub NC{alpha}} and the latter via interresidue two-bond {sup 2}H{sub NC{alpha}} couplings. The second step involves the identification of side-chain spin systems by 3D {sup 1}H-{sup 13}C-{sup 13}C-{sup 1}H correlated (HCCH-COSY) and 3D {sup 1}H-{sup 13}C-{sup 13}C-{sup 1}H total correlated (HCCH-TOCSY) spectroscopy, the latter making use of isotropic mixing of {sup 13}C magnetization to obtain relayed connectivities along the side chains. The authors were able to obtain complete {sup 1}H and {sup 13}C side-chain assignments for all residues, with the exception of 4 (out of a total of 15) lysine residues for which partial assignments were obtained.

• OSTI ID: 5706499
• Journal Information: Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 29:35; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550601* -- Medicine-- Unsealed Radionuclides in Diagnostics
62 RADIOLOGY AND NUCLEAR MEDICINE
BARYONS
CARBON 13
CARBON ISOTOPES
CHEMICAL SHIFT
ELEMENTARY PARTICLES
EVEN-ODD NUCLEI
FERMIONS
GROWTH FACTORS
HADRONS
ISOTOPES
LIGHT NUCLEI
LYMPHOKINES
MAGNETIC RESONANCE
MITOGENS
MOLECULAR STRUCTURE
NUCLEAR MAGNETIC RESONANCE
NUCLEI
NUCLEONS
ORGANIC COMPOUNDS
OVERHAUSER EFFECT
PH VALUE
PROTEINS
PROTONS
RESONANCE
STABLE ISOTOPES