RGD Reference Report - Essential cysteine residues of the type IIa Na+/Pi cotransporter. - Rat Genome Database

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Essential cysteine residues of the type IIa Na+/Pi cotransporter.

Authors: Kohler, K  Forster, IC  Stange, G  Biber, J  Murer, H 
Citation: Kohler K, etal., Pflugers Arch. 2003 May;446(2):203-10. Epub 2003 Mar 26.
RGD ID: 7243150
Pubmed: (View Article at PubMed) PMID:12739158
DOI: Full-text: DOI:10.1007/s00424-003-1039-6

The rat renal Na(+)/P(i) cotransporter (NaP(i)-IIa) contains 12 native cysteines. When individually replaced by a serine, none appears essential for proper expression and function. Nevertheless, the formation of one essential cysteine bridge (C5/C6), together with a postulated second bridge, is necessary. To determine the minimum cysteine residues required for functional NaP(i)-IIa, with the goal of generating a Cys-less backbone for structure-function studies, mutants were constructed in which multiple endogenous cysteines were replaced by serines in different combinations. In Xenopus oocytes, most mutants were functional, except those where cysteine pairs C4/C9, C4/C12 or C9/C12 were simultaneously deleted. This suggested that one of these pairs could form the second cysteine bridge essential for expression and/or protein function. Up to eight cysteines could therefore be removed to give a functional Cys-reduced NaP(i)-IIa with activity and kinetics comparable to the wild-type (WT). This construct, like all intermediate mutants and the WT, was insensitive to cysteine-modifying methanethiosulfonate (MTS) reagents. Moreover, by introducing a novel cysteine into the Cys-reduced NaP(i)-IIa at a site functionally important in the WT (Ser-460), the loss of transport function reported for mutant S460C, after exposure to MTS reagents, was recapitulated. This confirmed that the MTS reagent site of action was Cys-460 and that modification of native cysteines does not contribute to S460C behavior.

Gene Ontology Annotations    

Biological Process

Objects Annotated

Genes (Rattus norvegicus)
Slc34a1  (solute carrier family 34 member 1)

Additional Information