Time-resolved magnetic resonance spectroscopic study of freeze-trapped yeast.
Rapidly changing metabolic events in actively respiring yeast under strictly physiological conditions were approached by freeze-trapping analysis with 31P magnetic resonance spectroscopy. A 50% wet weight/volume suspension of Saccharomyces cerevisiae was freeze-trapped with 20% ethylene glycol and 5% methanol as an antifreezing agent, and with 3 mM creatine phosphate as an external standard. A phosphorus spectrum of the freeze-trapped yeast was measured for 20 min at -18 degrees C at 202.46 MHz, since energy-related phosphate compounds in yeast were stable at least for 20 min at the temperature. The time scale, defined as (time required for NMR measurement)/(time for trapping), was expanded by the freeze-trapping analysis by 67-times, since the time for trapping was 0.3 min. The use of creatine phosphate as an external standard enabled exact quantification of phosphorus resonances. 31P-NMR study of freeze-trapped yeast affords a well time-resolved and highly sensitive method to study phosphate metabolism.
Tanaka
A
A
Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia 19104.
Chance
B
B
Wehrli
S
S
D000255:Adenosine Triphosphate / Q000378:metabolism; D000332:Aerobiosis; D000693:Anaerobiosis; D004734:Energy Metabolism; D019855:Ethylene Glycol; D005026:Ethylene Glycols; D005615:Freezing; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D000432:Methanol; D010710:Phosphates / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*
D005026:Ethylene Glycols; D010710:Phosphates; D000255:Adenosine Triphosphate; D019855:Ethylene Glycol; D000432:Methanol