The Journal of Cell Biology, Vol 101, 1281-1287, Copyright © 1985 by The Rockefeller University Press
Organic anions stabilize the reactivated motility of sperm flagella and the latency of dynein 1 ATPase activity
BH Gibbons, WJ Tang and IR Gibbons
Substitution of any of a variety of organic anions, including acetate,
propionate, lactate, gluconate, and succinate, for chloride in the
reactivation medium improves the motility of demembranated sperm of
Tripneustes gratilla. At the optimum concentration of 0.20 N, all of these
anions improve the duration of motility, with lactate and gluconate being
the best. The Michaelis constant for beat frequency (Kmf) is lower
(0.11-0.14 mM at 22 degrees C) in most of the organic anions than it is in
Cl- (0.20 mM), and the minimum ATP concentration required to support
oscillatory beating is reduced from 10 microM in chloride to 2 microM in
acetate, which together indicate a greater affinity of the axonemal ATPase
for MgATP2- in the organic anions media. The maximal beat frequency, fmax,
is as high as 42 Hz in 0.2 N succinate compared to 31 Hz in Cl-, whereas
the mean bend angle averages 2.8 rad in acetate compared to 2.4 rad in Cl-;
these values give a calculated average velocity of tubule sliding of
approximately 15 micron/s in acetate and succinate, which is approximately
30% greater than the value of 11 micron/s observed in chloride. The
reactivated sperm are sixfold more sensitive to vanadate inhibition in 0.2
M acetate than they are in 0.15 M Cl-. The specific ATPase activity of
soluble dynein 1, which increases more than 15-fold between 0 and 1.0 N
Cl-, undergoes only a twofold activation over the same range of organic
anion concentration, and, like the reactivated motility, is up to 50-fold
more sensitive to vanadate. This greater apparent mechanochemical
efficiency and the increased sensitivity to vanadate inhibition in the
organic anions suggest that they, unlike chloride, do not promote the
spontaneous dissociation of ADP and PO4(3-) from the dynein-ADP-PO4 kinetic
intermediate in the dynein crossbridge cycle. The use of organic anion
media may lead to significant improvements in reactivation of other motile
and transport systems.
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