Cloning and activity of a novel α-latrotoxin from red-back spider venom

Graudins, A; Little, MJ; Pineda, SS; Hains, PG; King, GF; Broady, KW; Nicholson, GM

Cloning and activity of a novel α-latrotoxin from red-back spider venom

Graudins, A

Little, MJ Pineda, SS Hains, PG King, GF Broady, KW Nicholson, GM

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Publication Type:: Journal Article
Citation:: Biochemical Pharmacology, 2012, 83 (1), pp. 170 - 183
Issue Date:: 2012-01-01

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Field	Value	Language
dc.contributor.author	Graudins, A https://orcid.org/0000-0002-0310-3983	en_US
dc.contributor.author	Little, MJ	en_US
dc.contributor.author	Pineda, SS	en_US
dc.contributor.author	Hains, PG	en_US
dc.contributor.author	King, GF	en_US
dc.contributor.author	Broady, KW	en_US
dc.contributor.author	Nicholson, GM https://orcid.org/0000-0002-4277-4296	en_US
dc.date.available	2011-09-28	en_US
dc.date.issued	2012-01-01	en_US
dc.identifier.citation	Biochemical Pharmacology, 2012, 83 (1), pp. 170 - 183	en_US
dc.identifier.issn	0006-2952	en_US
dc.identifier.uri	http://hdl.handle.net/10453/18571
dc.description.abstract	The venom of the European black widow spider Latrodectus tredecimguttatus (Theridiidae) contains several high molecular mass (110-140 kDa) neurotoxins that induce neurotransmitter exocytosis. These include a vertebrate-specific α-latrotoxin (α-LTX-Lt1a) responsible for the clinical symptoms of latrodectism and numerous insect-specific latroinsectoxins (LITs). In contrast, little is known about the expression of these toxins in other Latrodectus species despite the fact that envenomation by these spiders induces a similar clinical syndrome. Here we report highly conserved α-LTX, α-LIT and δ-LIT sequence tags in Latrodectus mactans, Latrodectus hesperus and Latrodectus hasselti venoms using tandem mass spectrometry, following bioassay-guided separation of venoms by liquid chromatography. Despite this sequence similarity, we show that the anti-α-LTX monoclonal antibody 4C4.1, raised against α-LTX-Lt1a, fails to neutralize the neurotoxicity of all other Latrodectus venoms tested in an isolated chick biventer cervicis nerve-muscle bioassay. This suggests that there are important structural differences between α-LTXs in theridiid spider venoms. We therefore cloned and sequenced the α-LTX from the Australian red-back spider L. hasselti (α-LTX-Lh1a). The deduced amino acid sequence of the mature α-LTX-Lh1a comprises 1180 residues (∼132 kDa) with ∼93% sequence identity with α-LTX-Lt1a. α-LTX-Lh1a is composed of an N-terminal domain and a central region containing 22 ankyrin-like repeats. The presence of two furin cleavage sites, conserved with α-LTX-Lt1a, indicates that α-LTX-Lh1a is derived from the proteolytic cleavage of an N-terminal signal peptide and C-terminal propeptide region. However, we show that α-LTX-Lh1a has key substitutions in the 4C4.1 epitope that explains the lack of binding of the monoclonal antibody. © 2011 Elsevier Inc.	en_US
dc.relation.ispartof	Biochemical Pharmacology	en_US
dc.relation.isbasedon	10.1016/j.bcp.2011.09.024	en_US
dc.subject.classification	Pharmacology & Pharmacy	en_US
dc.subject.mesh	Muscle, Skeletal	en_US
dc.subject.mesh	Animals	en_US
dc.subject.mesh	Chickens	en_US
dc.subject.mesh	Black Widow Spider	en_US
dc.subject.mesh	Gryllidae	en_US
dc.subject.mesh	Spider Venoms	en_US
dc.subject.mesh	Cloning, Molecular	en_US
dc.subject.mesh	Toxicity Tests	en_US
dc.subject.mesh	Amino Acid Sequence	en_US
dc.subject.mesh	Base Sequence	en_US
dc.subject.mesh	Molecular Sequence Data	en_US
dc.subject.mesh	Female	en_US
dc.title	Cloning and activity of a novel α-latrotoxin from red-back spider venom	en_US
dc.type	Journal Article
utslib.citation.volume	1	en_US
utslib.citation.volume	83	en_US
utslib.for	111506 Toxicology (incl. Clinical Toxicology)	en_US
utslib.for	060199 Biochemistry and Cell Biology not elsewhere classified	en_US
utslib.for	0601 Biochemistry and Cell Biology	en_US
utslib.for	1115 Pharmacology and Pharmaceutical Sciences	en_US
pubs.embargo.period	Not known	en_US
pubs.organisational-group	/University of Technology Sydney
pubs.organisational-group	/University of Technology Sydney/Faculty of Science
pubs.organisational-group	/University of Technology Sydney/Faculty of Science/School of Life Sciences
pubs.organisational-group	/University of Technology Sydney/Strength - CHT - Health Technologies
utslib.copyright.status	open_access
pubs.issue	1	en_US
pubs.publication-status	Published	en_US
pubs.volume	83	en_US

Abstract:

The venom of the European black widow spider Latrodectus tredecimguttatus (Theridiidae) contains several high molecular mass (110-140 kDa) neurotoxins that induce neurotransmitter exocytosis. These include a vertebrate-specific α-latrotoxin (α-LTX-Lt1a) responsible for the clinical symptoms of latrodectism and numerous insect-specific latroinsectoxins (LITs). In contrast, little is known about the expression of these toxins in other Latrodectus species despite the fact that envenomation by these spiders induces a similar clinical syndrome. Here we report highly conserved α-LTX, α-LIT and δ-LIT sequence tags in Latrodectus mactans, Latrodectus hesperus and Latrodectus hasselti venoms using tandem mass spectrometry, following bioassay-guided separation of venoms by liquid chromatography. Despite this sequence similarity, we show that the anti-α-LTX monoclonal antibody 4C4.1, raised against α-LTX-Lt1a, fails to neutralize the neurotoxicity of all other Latrodectus venoms tested in an isolated chick biventer cervicis nerve-muscle bioassay. This suggests that there are important structural differences between α-LTXs in theridiid spider venoms. We therefore cloned and sequenced the α-LTX from the Australian red-back spider L. hasselti (α-LTX-Lh1a). The deduced amino acid sequence of the mature α-LTX-Lh1a comprises 1180 residues (∼132 kDa) with ∼93% sequence identity with α-LTX-Lt1a. α-LTX-Lh1a is composed of an N-terminal domain and a central region containing 22 ankyrin-like repeats. The presence of two furin cleavage sites, conserved with α-LTX-Lt1a, indicates that α-LTX-Lh1a is derived from the proteolytic cleavage of an N-terminal signal peptide and C-terminal propeptide region. However, we show that α-LTX-Lh1a has key substitutions in the 4C4.1 epitope that explains the lack of binding of the monoclonal antibody. © 2011 Elsevier Inc.

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http://hdl.handle.net/10453/18571