In Vitro Enzymatic Studies Reveal pH and Temperature Sensitive Properties of the CLIC Proteins.

Alghalayini, A; Hossain, KR; Moghaddasi, S; Turkewitz, DR; D'Amario, C; Wallach, M; Valenzuela, SM

In Vitro Enzymatic Studies Reveal pH and Temperature Sensitive Properties of the CLIC Proteins.

Alghalayini, A

Hossain, KR Moghaddasi, S Turkewitz, DR D'Amario, C Wallach, M Valenzuela, SM

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Publisher:: MDPI
Publication Type:: Journal Article
Citation:: Biomolecules, 2023, 13, (9), pp. 1394
Issue Date:: 2023-09-15

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Field	Value	Language
dc.contributor.author	Alghalayini, A https://orcid.org/0000-0002-2308-0256
dc.contributor.author	Hossain, KR
dc.contributor.author	Moghaddasi, S
dc.contributor.author	Turkewitz, DR
dc.contributor.author	D'Amario, C
dc.contributor.author	Wallach, M
dc.contributor.author	Valenzuela, SM
dc.date.accessioned	2024-01-15T04:21:40Z
dc.date.available	2023-09-10
dc.date.available	2024-01-15T04:21:40Z
dc.date.issued	2023-09-15
dc.identifier.citation	Biomolecules, 2023, 13, (9), pp. 1394
dc.identifier.issn	2218-273X
dc.identifier.issn	2218-273X
dc.identifier.uri	http://hdl.handle.net/10453/174442
dc.description.abstract	Chloride intracellular ion channel (CLIC) proteins exist as both soluble and integral membrane proteins, with CLIC1 capable of shifting between two distinct structural conformations. New evidence has emerged indicating that members of the CLIC family act as moonlighting proteins, referring to the ability of a single protein to carry out multiple functions. In addition to their ion channel activity, CLIC family members possess oxidoreductase enzymatic activity and share significant structural and sequence homology, along with varying overlaps in their tissue distribution and cellular localization. In this study, the 2-hydroxyethyl disulfide (HEDS) assay system was used to characterize kinetic properties, as well as the temperature and pH profiles of three CLIC protein family members (CLIC1, CLIC3, CLIC4). We also assessed the effects of the drugs rapamycin and amphotericin B, on the three CLIC proteins' enzymatic activity in the HEDS assay. Our results demonstrate CLIC1 to be highly heat-sensitive, with optimal enzymatic activity observed at neutral pH7 and at a temperature of 37 °C, while CLIC3 had higher oxidoreductase activity in more acidic pH5 and was found to be relatively heat stable. CLIC4, like CLIC1, was temperature sensitive with optimal enzymatic activity observed at 37 °C; however, it showed optimal activity in more alkaline conditions of pH8. Our current study demonstrates individual differences in the enzymatic activity between the three CLIC proteins, suggesting each CLIC protein is likely regulated in discrete ways, involving changes in the subcellular milieu and microenvironment.
dc.format	Electronic
dc.language	eng
dc.publisher	MDPI
dc.relation	http://purl.org/au-research/grants/arc/IH150100028
dc.relation.ispartof	Biomolecules
dc.relation.isbasedon	10.3390/biom13091394
dc.rights	info:eu-repo/semantics/openAccess
dc.subject	0601 Biochemistry and Cell Biology
dc.subject.classification	3101 Biochemistry and cell biology
dc.subject.classification	3102 Bioinformatics and computational biology
dc.subject.classification	3206 Medical biotechnology
dc.title	In Vitro Enzymatic Studies Reveal pH and Temperature Sensitive Properties of the CLIC Proteins.
dc.type	Journal Article
utslib.citation.volume	13
utslib.location.activity	Switzerland
utslib.for	0601 Biochemistry and Cell Biology
pubs.organisational-group	/University of Technology Sydney
pubs.organisational-group	/University of Technology Sydney/Faculty of Science
pubs.organisational-group	/University of Technology Sydney/Strength - CHT - Health Technologies
pubs.organisational-group	/University of Technology Sydney/Faculty of Science/School of Life Sciences
pubs.organisational-group	/University of Technology Sydney/Centre for Health Technologies (CHT)
utslib.copyright.status	open_access	*
dc.date.updated	2024-01-15T04:16:03Z
pubs.issue	9
pubs.publication-status	Published online
pubs.volume	13
utslib.citation.issue	9

Abstract:

Chloride intracellular ion channel (CLIC) proteins exist as both soluble and integral membrane proteins, with CLIC1 capable of shifting between two distinct structural conformations. New evidence has emerged indicating that members of the CLIC family act as moonlighting proteins, referring to the ability of a single protein to carry out multiple functions. In addition to their ion channel activity, CLIC family members possess oxidoreductase enzymatic activity and share significant structural and sequence homology, along with varying overlaps in their tissue distribution and cellular localization. In this study, the 2-hydroxyethyl disulfide (HEDS) assay system was used to characterize kinetic properties, as well as the temperature and pH profiles of three CLIC protein family members (CLIC1, CLIC3, CLIC4). We also assessed the effects of the drugs rapamycin and amphotericin B, on the three CLIC proteins' enzymatic activity in the HEDS assay. Our results demonstrate CLIC1 to be highly heat-sensitive, with optimal enzymatic activity observed at neutral pH7 and at a temperature of 37 °C, while CLIC3 had higher oxidoreductase activity in more acidic pH5 and was found to be relatively heat stable. CLIC4, like CLIC1, was temperature sensitive with optimal enzymatic activity observed at 37 °C; however, it showed optimal activity in more alkaline conditions of pH8. Our current study demonstrates individual differences in the enzymatic activity between the three CLIC proteins, suggesting each CLIC protein is likely regulated in discrete ways, involving changes in the subcellular milieu and microenvironment.

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http://hdl.handle.net/10453/174442