P-31 NUCLEAR MAGNETIC-RESONANCE STUDIES ON RELAXATION PARAMETERS AND LINE BROADENING OF INTRACELLULAR METABOLITES OF HELA-CELLS

被引:24
作者
EVANS, FE [1 ]
机构
[1] UNIV CALIF SAN DIEGO, DEPT CHEM, LA JOLLA, CA 92093 USA
基金
美国国家科学基金会; 美国国家卫生研究院;
关键词
D O I
10.1016/0003-9861(79)90009-2
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The 40 MHz 31P NMR spectrum of intact HeLa [human cervical cancer] cells contains 7 broad peaks with some detectable splittings. The linewidths were significantly broader than for those of cell-free systems such as cell extracts, indicating that the cellular environment is responsible for the unusual line broadening. Resolution of these peaks at 40 MHz is sufficient to make certain assignments and the relaxation parameters of some of the intracellular metabolites were measured. The spin-lattice relaxation times (T1) ranged from 0.3 s for ATP to about 3 s for Pi and monophosphate compounds. Nuclear Overhauser enhancements (NOE) were induced by proton irradiation with the possible exception of ATP. The relaxation parameters were compared to those of cell-free compounds; in all cases T1 and NOE were smaller for the intracellular metabolites. The relaxation parameters for ATP were affected the most. This behavior was mimicked with mixtures of cell-free metabolites containing paramagnetic ions. The larger change in both T1 and NOE of intracellular ATP could be accounted for by selective binding of paramagnetic ions. This phenomenon also explains some of the line broadening in the cell spectrum especially that of ATP. The spin-spin relaxation times (T2) of Pi and monophosphate compounds as measured by a pulse technique did not account for the observed linewidths. This is due to the presence of chemical shift envelopes arising from pH heterogeneity. All resonances were broader at 146 MHz because of the line broadening by paramagnetic ions and the presence of chemical shift envelopes. Other mechanisms of line broadening may also be significant. There was little difference in resolution of spectra at 40 and 146 MHz. Water proton linewidths and T2 values were measured for HeLa cells and for some minced tissue preparations. The water linewidth in tissue samples was broader than that in the cell suspension. The large linewidths in tissues arise mainly from chemical shift envelopes caused by magnetic field nonuniformity in the tissue samples. There appears to be a small chemical shift envelope from magnetic nonuniformity in HeLa cells as well. The 1H results on envelopes were extrapolated to 31P studies on cells and tissues. Possible methods for reducing linewidths arising from the various proposed broadening mechanisms were discussed.
引用
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页码:63 / 75
页数:13
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