Regions of Association between the α and the β Subunit of the Gastric H,K-ATPase

A binding and a yeast two-hybrid analysis were carried out on the gastric H,K-ATPase to determine interactive regions of the extracytoplasmic domains of the α and β subunits of this P type ATPase. Wheat germ agglutinin fractionation of fluorescein 5-maleimide-labeled tryptic fragments of detergent...

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Bibliographic Details
Published in:The Journal of biological chemistry 1998-05, Vol.273 (18), p.11075
Main Authors: Dominique Melle-Milovanovic, Marko Milovanovic, Sunil Nagpal, George Sachs, Jai Moo Shin
Format: Article
Language:English
Online Access:Get full text
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Summary:A binding and a yeast two-hybrid analysis were carried out on the gastric H,K-ATPase to determine interactive regions of the extracytoplasmic domains of the α and β subunits of this P type ATPase. Wheat germ agglutinin fractionation of fluorescein 5-maleimide-labeled tryptic fragments of detergent-solubilized H,K-ATPase showed that a fragment Leu 855 to Arg 922 of the α subunit was bound to the β subunit. The yeast two-hybrid system showed that the region containing only a part of the seventh transmembrane segment, the loop, and part of the eighth transmembrane segment was capable of giving positive interaction signals with the ectodomain of the β subunit. The sequence in the extracytoplasmic loop close to the eighth transmembrane segment, namely Arg 898 to Thr 928 , was identified as being the site of interaction using this method. We deduced that the sequence Arg 898 to Arg 922 in the α subunit has strong interaction with the extracytoplasmic domain of the β subunit. Again, using yeast two-hybrid analysis, two different sequences in the β subunit Gln 64 to Asn 130 and Ala 156 to Arg 188 were identified as association domains in the extracytoplasmic sequence of the β subunit. These data enable identification of major associative regions of the α-β subunits of the H,K-ATPase.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.273.18.11075