The biochemistry of serpin secretion : an investigation into the effects of mutant and corrective variants
The biochemistry of serpin secretion : an investigation into the effects of mutant and corrective variants
FENIB diseases are a distinct class of dementia pathologies, which show deposition of polymerised neuroserpin (P112, SERPIN1) in structures known as Collins’ bodies, which leads to the death of the synthesizing cells. Another feature of the diseases is the loss of neurons from well defined areas thought to arise from a deficiency of the neuroserpin.
There are four natural mutations in the neuroserpin gene that lead to these FENIB pathologies: Syracuse - S249P is the equivalent mutation to α1-AntitrypsinSilymama, which is at the commencement of the sheet-opening domain, Portland - S52R, H338R & G392E. These mutations are all within the shutter domain, a domain that is structurally important to the activity of all inhibitory serpins. All four mutants are autosomal dominant and lead to pathological phenotypes that can be ordered in severity: S49P<S52R<H338R<G392E.
We have examined the secretion of these four mutant forms of neuroserpin and compared it to that of the normal variant protein using the Xenopus oocyte system. The time-course of secretion for normal variant neuroserpin in comparison to that of another serpin, wildtype α1-antitrypsin was also studied.
58.1% (±6.4) of normal variant neuroserpin is secreted from oocytes microinjected with wildtype mRNA, slightly lower than seen in previous expression of M variant α1-antitrypsin (63%). All mutant variants reduce secretion: S49P 11.1% (±3.9) S52R 13.9% (±3.5), H338R 15.1%, G392E 9.9%, Y34S 2.2% (±2.1). Neuroserpin wildtype has a slower initial secretion than α1-AT which is maybe due to the polarised nature of secretion and could be directed by novel signalling motifs.
Using chemical and physical dissection, the intracellular location of the polymers visualised within this system was determined to be the ER. Endo H resistance was not inferred on oocyte extracts suggesting that accumulation occurs within a pre-Golgi compartment, with tunicamycin treatment showing that glycosylation is required for successful secretion but has no direct effect on polymerisation.
University of Southampton
Ravenhill, Neil A
f621445e-342a-4430-8c83-591314fd8bdb
2004
Ravenhill, Neil A
f621445e-342a-4430-8c83-591314fd8bdb
Ravenhill, Neil A
(2004)
The biochemistry of serpin secretion : an investigation into the effects of mutant and corrective variants.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
FENIB diseases are a distinct class of dementia pathologies, which show deposition of polymerised neuroserpin (P112, SERPIN1) in structures known as Collins’ bodies, which leads to the death of the synthesizing cells. Another feature of the diseases is the loss of neurons from well defined areas thought to arise from a deficiency of the neuroserpin.
There are four natural mutations in the neuroserpin gene that lead to these FENIB pathologies: Syracuse - S249P is the equivalent mutation to α1-AntitrypsinSilymama, which is at the commencement of the sheet-opening domain, Portland - S52R, H338R & G392E. These mutations are all within the shutter domain, a domain that is structurally important to the activity of all inhibitory serpins. All four mutants are autosomal dominant and lead to pathological phenotypes that can be ordered in severity: S49P<S52R<H338R<G392E.
We have examined the secretion of these four mutant forms of neuroserpin and compared it to that of the normal variant protein using the Xenopus oocyte system. The time-course of secretion for normal variant neuroserpin in comparison to that of another serpin, wildtype α1-antitrypsin was also studied.
58.1% (±6.4) of normal variant neuroserpin is secreted from oocytes microinjected with wildtype mRNA, slightly lower than seen in previous expression of M variant α1-antitrypsin (63%). All mutant variants reduce secretion: S49P 11.1% (±3.9) S52R 13.9% (±3.5), H338R 15.1%, G392E 9.9%, Y34S 2.2% (±2.1). Neuroserpin wildtype has a slower initial secretion than α1-AT which is maybe due to the polarised nature of secretion and could be directed by novel signalling motifs.
Using chemical and physical dissection, the intracellular location of the polymers visualised within this system was determined to be the ER. Endo H resistance was not inferred on oocyte extracts suggesting that accumulation occurs within a pre-Golgi compartment, with tunicamycin treatment showing that glycosylation is required for successful secretion but has no direct effect on polymerisation.
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Published date: 2004
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Local EPrints ID: 465497
URI: http://eprints.soton.ac.uk/id/eprint/465497
PURE UUID: cc2d6694-d025-4021-ae79-14801d3012e0
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Date deposited: 05 Jul 2022 01:27
Last modified: 16 Mar 2024 20:13
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Author:
Neil A Ravenhill
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