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), Vol. 29:35; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

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