Adenylate Kinase Activity in Ejaculated Bovine Sperm Flagella

P K Schoff,J Cheetham,H A Lardy

doi:10.1016/s0021-9258(18)83316-6

P K Schoff, J Cheetham + Show 1 more

Open Access

https://doi.org/10.1016/s0021-9258(18)83316-6

Copy DOI

Abstract

Adenylate kinase (ATP:AMP phosphotransferase, EC 2.7.4.3) activity was detected in the flagella of ejaculated bovine spermatozoa. This activity provided sufficient ATP to produce normal motility in cells permeabilized with digitonin and treated with 0.5 mM MgADP. In the presence of ADP, adenylate kinase activity was inhibited by P1,P5-di(adenosine 5')-pentaphosphate (Ap5A), an adenylate kinase-specific inhibitor, and motility was stopped. ATP-supported motility was not affected by Ap5A. Mitochondrial adenylate kinase activity allowed AMP to stimulate respiration in permeabilized sperm. Adenylate kinase activity in tail fragments was most active in a pH range from 7.6 to 8.4, and a similar pH sensitivity was observed for this enzyme activity in a hypotonic extract of whole sperm. The apparent km of adenylate kinase activity in permeabilized tail fragments was about 1.0 mM ADP in the direction of ATP synthesis. The fluctuation of nucleotide concentrations in normal and metabolically stimulated sperm suggested that adenylate kinase was most active when the cell was highly motile, although adenylate kinase activity did not appear to be coupled strictly with motility.

Highlights

Adenylate kinase (ATP:AMP phosphotransferase, EC 2.7.4.3) activitywas detected in the flagella of ejaculatedbovine spermatozoa.This activityprovided sufficient ATP to produce normalmotility in cellspermeabilized with digitonin and treated with 0.5 mM
The fluctuation of nucleotide concentrations in normaland metabolically stimulated sperm suggested that adenylate kinase wasmost active when the cell was highly motile, adenylate kinase activity did not appear to be coupledstrictly withmotility
Adenylate kinase is a ubiquitous enzyme that, by virtue of an equilibrium constant approaching 1, can produce either ADP or stoichiometric amounts of ATP and AMP depending on the concentrations of the three nucleotides

Summary

RESULTS

Reactivation of Motility in Ejaculated Bovine Sperm Per- cells may indicate that the inhibitor was degraded by sperm meabilized by Digitonin-Washed ejaculated sperm at 1 x 10' phosphatases. cells/ml became immotile within 0.5 min of exposure to 0.15 Production of ATP by Sperm Taik Using ADP as a SubmM digitonin (0.184 mg/ml). Activity of Adenylate Kinase in Intact Sperm-the concentrations of adenosine phosphates were constantand roughly equal over 30 min in motile sperm; treatment with 2.5 mM caffeine (Fig. 3B), which stimulated motility and generated a 2-fold increase in respiration (ll)d, rastically changed the adenosine phosphate distribution, decreasing ATP and increasing AMP. This shift in stimulated cells is consistent with the proposal that sperm tail adenylate kinase produces ATP from ADP, accumulating AMP. These cells retainedtheir responsiveness to AMP, indicating that theadenylate kinase activity was mitochondrial and notflagellar

DISCUSSION

B Caffeine

Findings

1.35 AFDiliPpin-wash