th  - 1988 - 51st ENC Conference

74
PH EFFECTS ON THE SOLUTION CONFORMATION OF
SHIKIMATE-3-PHOSPHATE: DETERMINATION BY NMR AND
DISTANCE GEOMETRY CALCULATIONS.
Stephen Castellino*, Gregory C. Leo and R. Doug Sammons
Monsanto Agricultural Company, A Unit of Monsanto Company,
800 N. Lindbergh Boulevard, St. Louis, Missouri 63167
The solution structure was determined for shikimate-3-phosphate (S3P), a cyclohexenyl
sugar derivative which is a metabolite in aromatic biosynthesis. Nuclear Overhauser effects (NOE)
and coupling constants were measured as a function of solution pH. The preferred conformation
found experimentally is the pseudo-axial chair with the 3-phosphate occupying an axial position.
This agrees with the conformation determined by force field calculations (MM-2). Comparisons of
experimental and theoretical three dimensional structures will be shown as a function of pH.
7s I
ASSIGNMENTS OF 31p AND IH RESONANCES IN OLIGONUCLEOTIDES BY TWO DIMENSIONAL
HETERONUCLEAR HARTMANN-HAHN SPECTROSCOPY. M.G. Zagorski*, M.W. Kalnik, X.Gao,
D. Norman, and M. Kouchakdjian, Department of Biochemistry and Molecular
Biophysics, College of Physicians and Surgeons, Columbia University, New York,
New York 10032, USA.
A modified he~eronuc~ar 2D shift correlation experiment is demonstrated
for assignment of H and P resonances in a variety of DNA olig~ucleotides
ranging in size from a self-complementar~1octamer (seven unique P atoms) up to
a complementary nonamer (sixteen unique J~P atoms). The modified pulse scheme
incorporates a MLEV-17 composite pulse scheme for achieving net magnetization
transfer among protons via a homonuclear Hartmann-Hahn (HOHAHA) type cross
polarization. This net homonuclear magnetization transfer is then relayed to
the --P atoms via an INEPT sequence and consequently recorded and processed to
provide high quality pure phase absorption spectra via the hypercomplex method.
Despite the relatively broad lin~ of these large DNA sequences, we were able to
unambiguously assign all of the -~P resonances and many of the C-I', C-2', C-4'
and C-5' sugar protons. These assignments were greatly facilitated by the
presence of rel~ cross peaks spanning 4 and 5 bonds from the C-I' and C-2'
protons to the --P nuclei. This relayed system removed many ambiguities present
from overlap of the C-3', C-4' and C-5' resonances. Although the experiment
described here does not provide the gains in sensitivity of the heteronuclear
multiple quantum coherence (indirect) experiments, it clearly provides an order
of magnitude improvement in sensitivity and resolution over ~her currently
available methods for (direct) detecting the less sensitive --P nuclei.
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