th  - 1988 - 51st ENC Conference

118 l APPLICATION OF N-H HETERONUCLEAR CORRELATION
SPECTROSCOPY TO SEVERAL 15N ENRICHED PROTEINS
Ed S. Mooberry* , Brian J. Stockman, Bin Yuan, Byung Ha Oh, and John L. Markley
National Magnetic Resonance Facility at Madison and Department of Biochemistry,
College of Agricultural and Life Sciences, University of Wisconsln-Madlson, Madison,
WI 53706
Several applications of N-H heteronuclear correlation spectroscopy (HETCOR) are
described for 15N enriched proteins. An experimental arrangement is shown for
obtaining 15N decoupled spectra. The proton chemical shift is removed from the
nitrogen dimension by using time proportional phase incrementation. Examples are
shown for [95% ul 15N]Anabaena 7120 flavodoxin (I), [95X ul 15N]Anabaena 7120
ferredoxln and [95% 15N-Leu]staphylococcal nuclease . The HETCOR spectra were
obtained by using Bruker reverse electronics with the new 451MHz IF frequency and a
5--- reverse probe. Water elimination was accomplished by solvent presaturatlon for
the flavodoxin and ferredoxin spectra. For nuclease, the spln-echo sequence (without
decoupling) recently published by Sklenar and Bax was used (2).
I. B.J. Stockman, W.M. Westler, E.S. Mooberry, and J.L. Markley, Biochemlstry'27, 136-
142 (1988). 1
2. V. Sklenar and A. Bax, J. Magn. Res. 74, 469-479 (1987).
[Supported by NIH Grants RR02301, RR02781, and GM35976, NSF Grant PCM-845048, and
USDA Competitive Research Grant 85-CRCR-I-1598.]
119 I 13C LABELING AND HIGH RESOLUTION 1H 2-D NMR: MAKING
UNNATURAL ESTERS STAND UP AND BE COUNTED
G.L. Helms~ W.P. Niemczura and R.E. Moore; Dept. of Chemistry,
University of Hawaii, Honolulu, HI. 96822
Polyhydroxylated natural products which also contain ester func-
tionalities can be formidable structure problems. It is often
necessary to peracetylate these compounds to alleviate solubility
problems or to increase spectral dispersion. Unfortunately the
sites of natural esterification now become indistinguishable from
those of the introduced esters. Ace~ylg~io ~ with 1,1'-'~C acetic
anhydride yields products in which -J --C--H couplings label the
sites of the introduced esters. These small couplings (I-4 Hz)
can be visualized even in crowded spectral regions by using
homonuclear H 2-D J Resolved or high resolution phase sensitive
COSY spectra. In both cases the heteronuclear coupling leads to a
splitting of the cross peak multiplet patterns in the F2 dim-
ension. This splitting not only indicates the location of t~e
introduced esters but also allows the determination of the-J
heteronuclear coupling constant. Examples taken from our work on
novel cyclic peptides and cyclodextrins will be presented.
158