Selective detection of mobile amino acid resonances of chloroplast ATP synthase by 1H spin-echo NMR

Wayne Frasch, W. D. Frasch

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

High-resolution proton spin-echo NMR spectra have been recorded for the solubilized chloroplast coupling factor 1 (CF1). A Hahn spin-echo sequence has been used to suppress the intense unresolved background signal that arises from protons in relatively structured regions of the enzyme, thereby permitting selective detection of the relatively small subset of protons which comprise the more mobile resonances on the enzyme. The 18-ms spin-echo spectrum consists of resonances which arise from approximately 120 protons (about 20 amino acids) per molecule of CF1. Characteristic line widths are in the range 1-10 Hz (T2 in the range 30-300 ms), which indicates a reorientational correlation time about 102 times shorter than that of the majority of proton resonances on CF1. Several peaks exhibit resolved multiplet structures and show J modulation in spin-echo spectra obtained at long τ. Specific resonances due to aspartate β-methylene protons, glutamate γ-methylene protons, lysine ε-methylene protons, and tyrosine aromatic protons have been assigned on the basis of (1) studies of the effects of selective covalent modification of the enzyme, (2) comparisons of observed chemical shifts with those of short reference peptides in aqueous media, and (3) spin-coupled multiplet structure. Chemical shifts of resonances assigned by covalent modification techniques correspond closely to values characteristic of aqueous media. Quantitative measurements indicate involvement of two to three lysines, one to two aspartates, one glutamate, and zero cysteine. The 18-ms spectra also contain resolved intensity from α-methine backbone protons in the region 3.9-4.6 ppm, suggesting the involvement of one or more random-coil peptide segments which extend into the aqueous phase.

Original languageEnglish (US)
Pages (from-to)5449-5454
Number of pages6
JournalBiochemistry
Volume24
Issue number20
StatePublished - 1985
Externally publishedYes

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Chloroplast Proton-Translocating ATPases
Protons
Nuclear magnetic resonance
Amino Acids
Chemical shift
Aspartic Acid
Lysine
Glutamic Acid
Enzymes
Peptides
Linewidth
Cysteine
Tyrosine

ASJC Scopus subject areas

  • Biochemistry

Cite this

Selective detection of mobile amino acid resonances of chloroplast ATP synthase by 1H spin-echo NMR. / Frasch, Wayne; Frasch, W. D.

In: Biochemistry, Vol. 24, No. 20, 1985, p. 5449-5454.

Research output: Contribution to journalArticle

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