Hexose-specificity of hexokinase and ADP-dependence of pyruvate kinase play important roles in the control of monosaccharide utilization in freshly diluted boar spermatozoa.
Incubation of boar sperm from fresh ejaculates in a minimal medium with 10 mM glucose induced a fast and intense activation of glycolysis, as indicated by the observed increases in the intracellular levels of glucose 6-phosphate (G 6-P) and ATP and the rate of formation of extracellular L-lactate. The effect of glucose was much more intense than that induced by fructose, sorbitol, and mannose. The greater utilization of glucose was related to a much greater sensitivity to hexokinase when compared with the other monosaccharides. Thus, the presence of 0.5 mM glucose induced total hexokinase activity in supernatants from sperm extracts of 1.7 +/- 0.1 mIU/mg protein, while the same concentration of both fructose, mannose, and sorbitol induced total hexokinase activity from 0.3 +/- 0.1 mIU/mg protein to 0.60 +/- 1 mIU/mg protein. Kinetic analysis of the total pyruvate kinase activity indicated that this activity was greatly dependent on the presence of ADP and also showed a great affinity for PEP, with an estimated Km in supernatants of 0.15-0.20 mM. Immunological location of proteins closely related to glycolysis, like GLUT-3 hexose transporter and hexokinase-I, indicated that these proteins showed the trend to be distributed around or in the cellular membranes of both head and midpiece in a grouped manner. We conclude that glycolysis is regulated by both the specific availability of a concrete sugar and the internal equilibrium between ATP and ADP levels. Furthermore, localization of proteins involved in the control of monosaccharide uptake and phosphorylation suggests that glycolysis starts at concrete points in the boar-sperm surface.
Medrano
Antonio
A
Department Animal Medicine and Surgery, Unit of Animal Reproduction, School of Veterinary Medicine, Autonomous University of Barcelona, Bellaterra, Spain.
García-Gil
Núria
N
Ramió
Laura
L
Montserrat Rivera
Maria
M
Fernández-Novell
Josep Maria
JM
Ramírez
Alfredo
A
Peña
Alejandro
A
Dolors Briz
Maria
M
Pinart
Elisabet
E
Concha
Ilona I
II
Bonet
Sergi
S
Rigau
Teresa
T
Rodríguez-Gil
Joan E
JE
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002245:Carbon Dioxide / Q000378:metabolism; D002457:Cell Extracts; D002470:Cell Survival / Q000187:drug effects; D004542:Ejaculation; D051274:Glucose Transporter Type 3 / Q000378:metabolism; D019298:Glucose-6-Phosphate / Q000378:metabolism; D006003:Glycogen / Q000378:metabolism; D006593:Hexokinase / Q000378:metabolism*; D006601:Hexoses / Q000378:metabolism* / Q000494:pharmacology; D007201:Indicator Dilution Techniques; D007700:Kinetics; D019344:Lactic Acid / Q000096:biosynthesis; D008297:Male; D046529:Microscopy, Electron, Transmission; D019000:Phosphotyrosine / Q000378:metabolism; D011770:Pyruvate Kinase / Q000378:metabolism*; D013076:Sperm Count; D013094:Spermatozoa / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D013379:Substrate Specificity; D013552:Swine / Q000378:metabolism*
D002457:Cell Extracts; D051274:Glucose Transporter Type 3; D006601:Hexoses; D002245:Carbon Dioxide; D019000:Phosphotyrosine; D019344:Lactic Acid; D019298:Glucose-6-Phosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D006003:Glycogen; D006593:Hexokinase; D011770:Pyruvate Kinase