Functionally Important Residues in the Predicted 3rd Transmembrane Domain of the Type IIa Sodium-phosphate Cotransporter (NaPi-IIa)

Functionally Important Residues in the Predicted 3rd Transmembrane Domain of the Type IIa... The type IIa Na+/Pi, cotransporter (NaPi-IIa) mediates electrogenic transport of three Na+ and one divalent Pi ion (and one net positive charge) across the cell membrane. Sequence comparison of electrogenic NaPi-IIa and IIb isoforms with the electroneutral NaPi-IIc isoform pointed to the third transmembrane domain (TMD-3) as a possibly significant determinant of substrate binding. To elucidate the role of TMD-3 in the topology and mechanism underlying NaPi-IIa function we subjected it to cysteine scanning mutagenesis. The constructs were expressed in Xenopus oocytes and Pi transport kinetics were assayed by electrophysiology and radiotracer uptake. Cys substitution resulted in only marginally altered kinetics of Pi transport in those mutants providing sufficient current for analysis. Only one site, at the extracellular end of TMD-3, appeared to be accessible to methanethiosulfonate reagents. However, additional mutations carried out at D224 (replaced by E, G or N) and N227 (replaced by D or Q) resulted in markedly altered voltage and substrate dependencies of the Pi-dependent currents. Replacing Asp-224 (highly conserved in electrogenic a and b isoforms) with Gly (the residue found in the electroneutral c isoform) resulted in a mutant that mediated electroneutral Na+-dependent Pi transport. Since electrogenic NaPi-II transports 3 Na+/transport cycle, whereas electroneutral NaPi-IIc only transports 2, we speculate that this loss of electrogenicity might result from the loss of one of the three Na+ binding sites in NaPi-IIa. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png The Journal of Membrane Biology Springer Journals

Functionally Important Residues in the Predicted 3rd Transmembrane Domain of the Type IIa Sodium-phosphate Cotransporter (NaPi-IIa)

Loading next page...
 
/lp/springer_journal/functionally-important-residues-in-the-predicted-3rd-transmembrane-7YVPwKWNM5
Publisher
Springer-Verlag
Copyright
Copyright © 2005 by Springer Science+Business Media, Inc.
Subject
Life Sciences; Human Physiology; Biochemistry, general
ISSN
0022-2631
eISSN
1432-1424
D.O.I.
10.1007/s00232-005-0796-x
Publisher site
See Article on Publisher Site

References

You’re reading a free preview. Subscribe to read the entire article.


DeepDyve is your
personal research library

It’s your single place to instantly
discover and read the research
that matters to you.

Enjoy affordable access to
over 12 million articles from more than
10,000 peer-reviewed journals.

All for just $49/month

Explore the DeepDyve Library

Unlimited reading

Read as many articles as you need. Full articles with original layout, charts and figures. Read online, from anywhere.

Stay up to date

Keep up with your field with Personalized Recommendations and Follow Journals to get automatic updates.

Organize your research

It’s easy to organize your research with our built-in tools.

Your journals are on DeepDyve

Read from thousands of the leading scholarly journals from SpringerNature, Elsevier, Wiley-Blackwell, Oxford University Press and more.

All the latest content is available, no embargo periods.

See the journals in your area

Monthly Plan

  • Read unlimited articles
  • Personalized recommendations
  • No expiration
  • Print 20 pages per month
  • 20% off on PDF purchases
  • Organize your research
  • Get updates on your journals and topic searches

$49/month

Start Free Trial

14-day Free Trial

Best Deal — 39% off

Annual Plan

  • All the features of the Professional Plan, but for 39% off!
  • Billed annually
  • No expiration
  • For the normal price of 10 articles elsewhere, you get one full year of unlimited access to articles.

$588

$360/year

billed annually
Start Free Trial

14-day Free Trial