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Sex steroid binding proteins in fish


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Hobby, AC 2000 , 'Sex steroid binding proteins in fish', PhD thesis, University of Tasmania.

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A sex steroid binding protein (SBP) binding E2 with high affinity has been detected in the
pleuronectid greenback flounder (Rhombosolea tapirina), the sparids black bream
(Acanthopagrus butcheri) and snapper (Pagrus auratus), and its presence has been
confirmed in the salmonid rainbow trout (Oncorhynchus mykiss). SBP binding
characteristics were measured using a hot saturation assay for trout, bream and snapper and
a cold saturation assay for flounder. Bound and unbound steroid were separated by
incubation with dextran-coated charcoal (DCC). Affinity for E2 was highest in trout (kD=0.44 nM), followed by bream (kD=3.39 nM) and snapper (kD=10.7 nM). The lowest affinity was found in flounder (kD=84.7 nM). Binding capacity, however, was greatest in flounder (Bmax=164 nM), followed by trout (Bmax=92 nM), and then bream and snapper (Bmax=50 and 39 nM, respectively). Binding of E2 to SBP had a very rapid rate of
association, and most dissociation occurred within 5 min.
To confirm that the plasma protein measured here was SBP, the relative binding affinities
of SBP for a range of steroids was measured. In trout, bream and snapper, SBP bound E2
with the highest affinity, followed by T. In contrast, flounder SBP bound T more than
twice as strongly as E2. The rank orders of affinity of binding indicate the importance of an
unhindered 17β-hydroxyl group, and a 3-hydroxyl or 3-ketone group for high affinity
binding to SBP. These requirements for high affinity binding are present in most animals
possessing SBP and indicate conservation of the SBP molecule through evolution. The presence of seasonal changes in SBP binding characteristics was investigated in female
trout, bream and flounder. The binding capacity of trout and bream SBP was significantly
greater in vitellogenic than non-reproductive stage fish. A decrease in binding affinity was
found in male trout injected with E2 compared to control fish. This difference was
removed by partial purification of SBP by gel filtration, and may have resulted from
competitive inhibition of E2 binding by vitellogenin. No differences in flounder SBP were
The effect of short term confinement stress on SBP binding characteristics was examined
in female bream, and trout. Confinement of bream for 6 h (but not 1 or 24 h) after capture
significantly reduced the binding capacity of SBP. No significant differences in binding
characteristics were detected in trout after 5 h confinement, or at 3 and 6 h after treatment
with either cortisol or saline, although binding capacity tended to be lower in fish treated
with cortisol rather than saline at 6 h post-injection. Relative binding studies indicated that
plasma cortisol at concentrations 100x or more greater than plasma E2 may displace E2
from SBP in bream. Physiological levels of cortisol did not displace E2 from SBP in trout.
The results presented here predominantly support the idea of SBP as a steroid reservoir,
protecting and transporting E2 and/or T to target tissues. However, binding characteristics
of SBP differ between species and changes of SBP with reproductive development are not
consistent. This suggests that the either the role of SBP in reproduction may not be
profound, it may be important in a way not investigated in the present study, or SBP may
have a role that differs in some way between different species.

Item Type: Thesis - PhD
Authors/Creators:Hobby, AC
Keywords: Fishes, Steroid-binding proteins
Copyright Holders: The Author
Copyright Information:

Copyright 2000 the Author - The University is continuing to endeavour to trace the copyright
owner(s) and in the meantime this item has been reproduced here in good faith. We
would be pleased to hear from the copyright owner(s).

Additional Information:

Thesis (PhD )--University of Tasmania, 2000. Includes bibliographical references

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