Measurement of the Dexree of Coupled Isotopic Enrichment of Different Positions in an Antibiotic Peptide by NMR

A. F. Miller, L. A. Egan, C. A. Townsend

Research output: Contribution to journalArticlepeer-review


An experimental strategy for determining the extent to which multiply isotopically labeled fragments are incorporated intact into relatively complicated compounds of interest is presented. The NMR methods employed are based on isotope-filtered one-dimensional spectra and difference HSQC spectra incorporating a spin echo designed to report on the presence of a second NMR active isotope at a coupled site. They supplement existing methods for determining the extent of isotopic incorporation at individual sites to reveal whether two coupled labeled sites in a precursor are incorporated as an intact unit into products. The methods described also circumvent 1H signal overlap and distinguish between the effects of different nitrogens coupled to individual carbons. The somewhat complicated case of valclavam illustrates the method's utility in measuring the J coupling constants between 13C and nearby sites that are only fractionally labeled with 15N, and measuring the fraction of molecules in which 13C is coupled to 15N, at each of several sites. The 15N of [2-13C, 15N] -labeled glycine is found to be incorporated into all three N positions of valclavam but most heavily into the N11 position. Specifically, 15N and 13C are incorporated into the N11 and C10 positions together as an 15N13C fragment approximately 8% of the time, whereas 15N is incorporated largely independently at the other positions,

Original languageEnglish
Pages (from-to)120-131
Number of pages12
JournalJournal of Magnetic Resonance
Issue number1
StatePublished - Mar 1997

Bibliographical note

Funding Information:
We thank Professor Lewis Kay for generously providing a pulse sequence for CT-HSQC and Cambridge Isotope Labs for the gift of some 15N threonine. A.-F.M. gratefully acknowledges financial support from the N.S.F. MCB-9418181. Thanks are given to the donors of the Petroleum Research Fund, administered by the ACS, for partial support of this research under ACS-PRF 28379-G4. C.A.T. is pleased to thank the N.I.H. for financial support (AI14937). We thank the N.I.H. (RR 06468) for generous financial aid in acquiring the NMR spectrometer used.

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Nuclear and High Energy Physics
  • Condensed Matter Physics


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