Secondary structure and side-chain sup 1 H and sup 13 C resonance assignments of calmodulin in solution by heteronuclear multidimensional NMR spectroscopy

Journal Article · Tue Sep 24 00:00:00 EDT 1991 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00102a013· OSTI ID:5601571

Ikura, Mitsuhiko; Spera, S; Barbato, G; Kay, L E; Bax, A ^[1]; Krinks, M ^[2]

---

1. National Inst. of Health, Bethesda, MD (United States)
2. National Cancer Inst., Bethesda, MD (United States)

Heteronuclear 2D and 3D NMR experiments were carried out on recombinant Drosophila calmodulin (CaM), a protein of 148 residues and with molecular mass of 16.7 kDa, that is uniformly labeled with {sup 15}N and {sup 13}C to a level of > 95%. Nearly complete {sup 1}H and {sup 13}C side-chain assignments for all amino acid residues are obtained by using the 3D HCCH-COSY and HCCH-TOCSY experiments that rely on large heteronuclear one-bond scalar couplings to transfer magnetization and establish through-bond connectivities. The secondary structure of this protein in solution has been elucidated by a qualitative interpretation of nuclear Overhauser effects, hydrogen exchange data, and {sup 3}J{sub HNH{alpha}} coupling constants. A clear correlation between the {sup 13}C{alpha} chemical shift and secondary structure is found. The secondary structure in the two globular domains of Drosophila CaM in solution is essentially identical with that of the X-ray crystal structure of mammalian CaM which consists of two pairs of a helix-loop-helix motif in each globular domain. The existence of a short antiparallel {beta}-sheet between the two loops in each domain has been confirmed. The eight {alpha}-helix segments identified from the NMR data are located at Glu-6 to Phe-19, thr-29 to Ser-38, Glu-45 to Glu-54, Phe-65 to Lys-77, Glu-82 to Asp-93, Ala-102 to Asn-111, Asp-118 to Glu-127, and Tyr-138 to Thr-146. Although the crystal structure has a long central helix from Phe-65 to Phe-92 that connects the two globular domains, NMR data indicate that residues Asp-78 to Ser-81 of this central helix adopt a nonhelical conformation with considerable flexibility.

• OSTI ID: 5601571
• Journal Information: Biochemistry; (United States), Vol. 30:38; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

62 RADIOLOGY AND NUCLEAR MEDICINE
CALMODULIN
NUCLEAR MAGNETIC RESONANCE
CARBON 13
CHEMICAL SHIFT
DROSOPHILA
MOLECULAR STRUCTURE
OVERHAUSER EFFECT
PROTONS
ANIMALS
ARTHROPODS
BARYONS
CARBON ISOTOPES
DIPTERA
ELEMENTARY PARTICLES
EVEN-ODD NUCLEI
FERMIONS
FLIES
FRUIT FLIES
HADRONS
INSECTS
INVERTEBRATES
ISOTOPES
LIGHT NUCLEI
MAGNETIC RESONANCE
NUCLEI
NUCLEONS
ORGANIC COMPOUNDS
PROTEINS
RESONANCE
STABLE ISOTOPES
550601* - Medicine- Unsealed Radionuclides in Diagnostics