Functional expression of gastric H,K-ATPase using the baculovirus expression system

Corné H.W. Klaassen; Tom J.F. Van Uem; Mariëlle P. De Moel; Godelieve L.J. De Caluwé; Herman G.P. Swarts; Jan Joep H.H.M. De Pont

doi:10.1016/0014-5793(93)80237-O

Functional expression of gastric H,K-ATPase using the baculovirus expression system

Corné H.W. Klaassen, Tom J.F. Van Uem, Mariëlle P. De Moel, Godelieve L.J. De Caluwé, Herman G.P. Swarts, Jan Joep H.H.M. De Pont^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

60 Citations (Scopus)

Abstract

A novel approach to construct a single recombinant baculovirus expressing two protein subunits simultaneously by replacing polyhedrin as well as p10 coding sequences is described. The recombinant baculovirus expressed the α- as well as the β-subunit of the gastric H.K-ATPase. Sf9 cells infected with this virus exhibited a K⁺- and SCH 28080-sensitive ATP-dependent phosphorylation capacity in purified Sf9 membranes similar to Natlve H,K-ATPase. This activity was not present in control membranes containing only one of the two H,K-ATPase subunits. We therefore conclude that both subunits are essential for the phosphorylation capacity of H.K-ATPase.

Original language	English
Pages (from-to)	277-282
Number of pages	6
Journal	FEBS Letters
Volume	329
Issue number	3
DOIs	https://doi.org/10.1016/0014-5793(93)80237-O
Publication status	Published - 30 Aug 1993
Externally published	Yes

Bibliographical note

Funding Information:
AcknowledgemenrsT. he authors wish to thank Dr. Gary Shull for his donation of H,K-ATPase cDNA clones, Dr. Just Vlak for the transfer vectors pAcDZ1 and pAcAS3, St9 cells and weld type AcNPV, Drs. Mtchael Caplan, John Forte and Annick Soumarmon for their antt-bodies, Dr. Bjijrn Wallmark for his gift of SCH 28080 and Dr. Wim De Grip for hts advice in the initial phase of this project This work was sponsored by the Netherlands Foundation for SctenttficR esearch (NWO) under Grant 900-522-086.

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abstract = "A novel approach to construct a single recombinant baculovirus expressing two protein subunits simultaneously by replacing polyhedrin as well as p10 coding sequences is described. The recombinant baculovirus expressed the α- as well as the β-subunit of the gastric H.K-ATPase. Sf9 cells infected with this virus exhibited a K+- and SCH 28080-sensitive ATP-dependent phosphorylation capacity in purified Sf9 membranes similar to Natlve H,K-ATPase. This activity was not present in control membranes containing only one of the two H,K-ATPase subunits. We therefore conclude that both subunits are essential for the phosphorylation capacity of H.K-ATPase.",

author = "Klaassen, {Corn{\'e} H.W.} and {Van Uem}, {Tom J.F.} and {De Moel}, {Mari{\"e}lle P.} and {De Caluw{\'e}}, {Godelieve L.J.} and Swarts, {Herman G.P.} and {De Pont}, {Jan Joep H.H.M.}",

note = "Funding Information: AcknowledgemenrsT. he authors wish to thank Dr. Gary Shull for his donation of H,K-ATPase cDNA clones, Dr. Just Vlak for the transfer vectors pAcDZ1 and pAcAS3, St9 cells and weld type AcNPV, Drs. Mtchael Caplan, John Forte and Annick Soumarmon for their antt-bodies, Dr. Bjijrn Wallmark for his gift of SCH 28080 and Dr. Wim De Grip for hts advice in the initial phase of this project This work was sponsored by the Netherlands Foundation for SctenttficR esearch (NWO) under Grant 900-522-086.",

year = "1993",

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doi = "10.1016/0014-5793(93)80237-O",

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AU - Klaassen, Corné H.W.

AU - Van Uem, Tom J.F.

AU - De Moel, Mariëlle P.

AU - De Caluwé, Godelieve L.J.

AU - Swarts, Herman G.P.

AU - De Pont, Jan Joep H.H.M.

N1 - Funding Information: AcknowledgemenrsT. he authors wish to thank Dr. Gary Shull for his donation of H,K-ATPase cDNA clones, Dr. Just Vlak for the transfer vectors pAcDZ1 and pAcAS3, St9 cells and weld type AcNPV, Drs. Mtchael Caplan, John Forte and Annick Soumarmon for their antt-bodies, Dr. Bjijrn Wallmark for his gift of SCH 28080 and Dr. Wim De Grip for hts advice in the initial phase of this project This work was sponsored by the Netherlands Foundation for SctenttficR esearch (NWO) under Grant 900-522-086.

PY - 1993/8/30

Y1 - 1993/8/30

N2 - A novel approach to construct a single recombinant baculovirus expressing two protein subunits simultaneously by replacing polyhedrin as well as p10 coding sequences is described. The recombinant baculovirus expressed the α- as well as the β-subunit of the gastric H.K-ATPase. Sf9 cells infected with this virus exhibited a K+- and SCH 28080-sensitive ATP-dependent phosphorylation capacity in purified Sf9 membranes similar to Natlve H,K-ATPase. This activity was not present in control membranes containing only one of the two H,K-ATPase subunits. We therefore conclude that both subunits are essential for the phosphorylation capacity of H.K-ATPase.

AB - A novel approach to construct a single recombinant baculovirus expressing two protein subunits simultaneously by replacing polyhedrin as well as p10 coding sequences is described. The recombinant baculovirus expressed the α- as well as the β-subunit of the gastric H.K-ATPase. Sf9 cells infected with this virus exhibited a K+- and SCH 28080-sensitive ATP-dependent phosphorylation capacity in purified Sf9 membranes similar to Natlve H,K-ATPase. This activity was not present in control membranes containing only one of the two H,K-ATPase subunits. We therefore conclude that both subunits are essential for the phosphorylation capacity of H.K-ATPase.

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ER -

Functional expression of gastric H,K-ATPase using the baculovirus expression system

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