2-Deoxyglucose Transport and Phosphorylation by Bovine Sperm1

Abstract

Measurement of sugar transport in ejaculated bovine sperm is complicated by a small intracellular volume and a rapid rate of sugar uptake. However, initial rates of sugar uptake can be obtained using 2-deoxyglucose as a model substrate. Accumulation of 2-deoxyglucose involves both transport and phosphorylation and can be monitored accurately by measuring only intracellular 2-deoxyglucose-6-phosphate. Uptake of 2-deoxyglucose was linear for 10 seconds at 22°C and followed simple Michaelis-Menten kinetics with a K_m of 0.2 mM and a V_max of 2 µmole/h/10⁸ cells. Phloretin was a competitive and cytochalasin B a noncompetitive inhibitor of 2-deoxyglucose uptake and since neither compound inhibited hexokinase, transport is the rate determining step in the uptake of 2-deoxyglucose. Comparison of the inhibition kinetics of 2-deoxyglucose uptake and of sperm hexokinase using several sugars also supports this conclusion.