Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors.
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| Title: | Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors. |
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| Authors: | Anyatonwu, Georgia1, Khan, M. Tariq1, Schug, Zachary T.1, da Fonseca, Paula C. A.2, Morris, Edward P.2, Joseph, Suresh K.1 suresh.joseph@jefferson.edu |
| Source: | Journal of Biological Chemistry. 8/6/2010, Vol. 285 Issue 32, p25085-25093. 9p. 3 Diagrams, 4 Graphs. |
| Subjects: | Trypsin, Epitopes, Thiols, Inositol phosphates, Biochemistry |
| Abstract: | We have used limited trypsin digestion and reactivity with PEG-maleimides (MPEG) to study Ca2+-induced conformational changes of IP3Rs in their native membrane environment. We found that Ca2+ decreased the formation of the 95-kDa C-terminal tryptic fragment when detected by an Ab directed at a C-terminal epitope (CT-1) but not with an Ab recognizing a protected intraluminal epitope. This suggests that Ca2+ induces a conformational change in the IP3R that allows trypsin to cleave the C-terminal epitope. Half-maximal effects of Ca2+ were observed at ∼0.5 µM and was sensitive to inhibition by IP3. Ca2+ also stimulated the reaction of MPEG-5 with an endogenous thiol in the 95-kDa fragment. This effect was eliminated when six closely spaced cysteine residues proximal to the transmembrane domains were mutated (C2000S, C2008S, C2010S, C2043S, C2047S, and C2053S) or when the N-terminal suppressor domain (amino acids 1-225) was deleted. A cysteine substitution mutant introduced at the C-terminal residue (A2749C) was freely accessible to MPEG-5 or MPEG-20 in the absence of Ca2+. However, cysteine substitution mutants in the interior of the tail were poorly reactive with MPEG-5, although reactivity was enhanced by Ca2+. We conclude the following: a) that large conformational changes induced by Ca2+ can be detected in IP3Rs in situ; b) these changes may be driven by Ca2+ binding to the N-terminal suppressor domain and expose a group of closely spaced endogenous thiols in the channel domain; and c) that the C-terminal cytosol-exposed tail of the IP3R may be relatively inaccessible to regulatory proteins unless Ca2+ is present. [ABSTRACT FROM AUTHOR] |
| Copyright of Journal of Biological Chemistry is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.) | |
| Database: | Engineering Source |
| FullText | Text: Availability: 0 |
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| Header | DbId: egs DbLabel: Engineering Source An: 53446892 AccessLevel: 6 PubType: Academic Journal PubTypeId: academicJournal PreciseRelevancyScore: 0 |
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| Items | – Name: Title Label: Title Group: Ti Data: Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors. – Name: Author Label: Authors Group: Au Data: <searchLink fieldCode="AR" term="%22Anyatonwu%2C+Georgia%22">Anyatonwu, Georgia</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Khan%2C+M%2E+Tariq%22">Khan, M. Tariq</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Schug%2C+Zachary+T%2E%22">Schug, Zachary T.</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22da+Fonseca%2C+Paula+C%2E+A%2E%22">da Fonseca, Paula C. A.</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Morris%2C+Edward+P%2E%22">Morris, Edward P.</searchLink><relatesTo>2</relatesTo><br /><searchLink fieldCode="AR" term="%22Joseph%2C+Suresh+K%2E%22">Joseph, Suresh K.</searchLink><relatesTo>1</relatesTo><i> suresh.joseph@jefferson.edu</i> – Name: TitleSource Label: Source Group: Src Data: <searchLink fieldCode="JN" term="%22Journal+of+Biological+Chemistry%22">Journal of Biological Chemistry</searchLink>. 8/6/2010, Vol. 285 Issue 32, p25085-25093. 9p. 3 Diagrams, 4 Graphs. – Name: Subject Label: Subjects Group: Su Data: <searchLink fieldCode="DE" term="%22Trypsin%22">Trypsin</searchLink><br /><searchLink fieldCode="DE" term="%22Epitopes%22">Epitopes</searchLink><br /><searchLink fieldCode="DE" term="%22Thiols%22">Thiols</searchLink><br /><searchLink fieldCode="DE" term="%22Inositol+phosphates%22">Inositol phosphates</searchLink><br /><searchLink fieldCode="DE" term="%22Biochemistry%22">Biochemistry</searchLink> – Name: Abstract Label: Abstract Group: Ab Data: We have used limited trypsin digestion and reactivity with PEG-maleimides (MPEG) to study Ca2+-induced conformational changes of IP3Rs in their native membrane environment. We found that Ca2+ decreased the formation of the 95-kDa C-terminal tryptic fragment when detected by an Ab directed at a C-terminal epitope (CT-1) but not with an Ab recognizing a protected intraluminal epitope. This suggests that Ca2+ induces a conformational change in the IP3R that allows trypsin to cleave the C-terminal epitope. Half-maximal effects of Ca2+ were observed at ∼0.5 µM and was sensitive to inhibition by IP3. Ca2+ also stimulated the reaction of MPEG-5 with an endogenous thiol in the 95-kDa fragment. This effect was eliminated when six closely spaced cysteine residues proximal to the transmembrane domains were mutated (C2000S, C2008S, C2010S, C2043S, C2047S, and C2053S) or when the N-terminal suppressor domain (amino acids 1-225) was deleted. A cysteine substitution mutant introduced at the C-terminal residue (A2749C) was freely accessible to MPEG-5 or MPEG-20 in the absence of Ca2+. However, cysteine substitution mutants in the interior of the tail were poorly reactive with MPEG-5, although reactivity was enhanced by Ca2+. We conclude the following: a) that large conformational changes induced by Ca2+ can be detected in IP3Rs in situ; b) these changes may be driven by Ca2+ binding to the N-terminal suppressor domain and expose a group of closely spaced endogenous thiols in the channel domain; and c) that the C-terminal cytosol-exposed tail of the IP3R may be relatively inaccessible to regulatory proteins unless Ca2+ is present. [ABSTRACT FROM AUTHOR] – Name: AbstractSuppliedCopyright Label: Group: Ab Data: <i>Copyright of Journal of Biological Chemistry is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract.</i> (Copyright applies to all Abstracts.) |
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| RecordInfo | BibRecord: BibEntity: Identifiers: – Type: doi Value: 10.1074/jbc.M110.123208 Languages: – Code: eng Text: English PhysicalDescription: Pagination: PageCount: 9 StartPage: 25085 Subjects: – SubjectFull: Trypsin Type: general – SubjectFull: Epitopes Type: general – SubjectFull: Thiols Type: general – SubjectFull: Inositol phosphates Type: general – SubjectFull: Biochemistry Type: general Titles: – TitleFull: Calcium-dependent Conformational Changes in Inositol Trisphosphate Receptors. Type: main BibRelationships: HasContributorRelationships: – PersonEntity: Name: NameFull: Anyatonwu, Georgia – PersonEntity: Name: NameFull: Khan, M. Tariq – PersonEntity: Name: NameFull: Schug, Zachary T. – PersonEntity: Name: NameFull: da Fonseca, Paula C. A. – PersonEntity: Name: NameFull: Morris, Edward P. – PersonEntity: Name: NameFull: Joseph, Suresh K. IsPartOfRelationships: – BibEntity: Dates: – D: 06 M: 08 Text: 8/6/2010 Type: published Y: 2010 Identifiers: – Type: issn-print Value: 00219258 Numbering: – Type: volume Value: 285 – Type: issue Value: 32 Titles: – TitleFull: Journal of Biological Chemistry Type: main |
| ResultId | 1 |
