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Title
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1H NMR sequential resonance assignments, secondary structure, and global fold in solution of the major (trans-Pro43) form of bovine calbindin D9k.
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Authors
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J.Kördel,
S.Forsén,
W.J.Chazin.
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Ref.
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Biochemistry, 1989,
28,
7065-7074.
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PubMed id
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Abstract
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A wide range of two-dimensional 1H NMR experiments have been used to completely
assign the 500-MHz 1H NMR spectrum of recombinant Ca2+-saturated bovine
calbindin D9k (76 amino acids, Mr = 8500). In solution, calbindin D9k exists as
an equilibrium mixture of isoforms with trans (75%) and cis (25%) isomers of the
peptide bond at Pro43 [Chazin et al. (1989) Proc. Natl. Acad. Sci. U.S.A. 86,
2195-2198], which results in two sets of 1H NMR signals from approximately half
of the amino acids. The complete 1H NMR assignments for the major, trans-Pro43
isoform are presented here. By use of an integrated strategy for spin system
identification, 62 of the 76 spin systems could be assigned to the appropriate
residue type. Sequence-specific assignments were then obtained by the standard
method. Secondary structure elements were identified on the basis of networks of
sequential and medium-range nuclear Overhauser effects (NOEs), 3JHN alpha spin
coupling constants, and the location of slowly exchanging amide protons. Four
helical segments and a short beta-sheet between the two calcium binding loops
are found. These elements of secondary structure and a few additional long-range
NOEs provide the global fold. Good agreement is found between the solution and
crystal structures of the minor A form of bovine calbindin D9k and between the
solution structures of the minor A form of bovine calbindin D9k and intact
porcine calbindin D9k.
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