Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
2000-01-14
2003-03-18
Jones, W. Gary (Department: 1634)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C530S333000, C530S350000
Reexamination Certificate
active
06534275
ABSTRACT:
TECHNICAL FIELD
The present invention is in the field of molecular biology; more particularly, the present invention describes the nucleic acid and amino acid sequences of three novel cellubrevin homologs.
BACKGROUND ART
Cellubrevins are homologs of synaptobrevins, synaptic vesicle-associated membrane proteins (VAMPs). Synaptobrevin was first discovered in rat brain (Baumert et al (1989) Embo J 8:379-84) and initially thought to be limited to neuronal cells. Synaptobrevin is an integral membrane protein of 18 kDA (Ralston E et al (1994) J Biol Chem 269:15403-6) involved in the movement of vesicles from the plasmalemma of one cell, across the synapse, to the plasma membrane of the receptive neuron. This regulated vesicle trafficking pathway and the endocytotic process may be blocked by the highly specific action of clostridial neurotoxins which prevents neurotransmitter release by cleaving the synaptobrevin molecule. Synaptobrevins are now known to occur and function in constitutive vesicle trafficking pathways involving the receptor-mediated endocytotic and exocytotic pathways of many non-neuronal cell types.
Cellubrevins are 16 kDa proteins first found and investigated in rat cells and tissues (McMahon HT et al (1993) Nature 364:346-9). In vitro studies of various cellular membranes (Galli et al (1994) J Cell Biol 125:1015-24; Link et al (1993) J Biol Chem 268:18423-6) have shown that VAMPS including the cellubrevins are widely distributed and are important in membrane trafficking. They appear to participate in axon extension via exocytosis during development, in the release of neurotransmitters and modulatory peptides, and in endocytosis. Endocytotic vesicular transport includes such intracellular events as the fusions and fissions of the nuclear membrane, endoplasmic reticulum, Golgi apparatus, and various inclusion bodies such as peroxisomes or lysosomes.
Endocytotic processes appear to be universal in eukaryotic cells as diverse as yeast,
Caenorhabditis elegans
, Drosophila, and humans. The homologous proteins which direct the movement of vesicles within and between the cells of these organisms contain evolutionarily conserved domains. Generally, VAMPs have a three domain organization. The domains include a variable proline-rich, N-terminal sequence of 28 amino acids, a highly conserved central hydrophilic core of 69 amino acids, and a hydrophobic sequence of 23 amino acids presumed to be the membrane anchor.
As mentioned for synaptobrevin above, cellubrevins are sensitive to selective proteolysis by metalloendoproteases such as the zinc endoprotease which comprises the light chain of tetanus toxin. Experiments have shown that endosome fusion may continue even after specific cellubrevin cleavage through temperature- and ATP-dependent docking and fusion processes involving N-ethylmaleimide-sensitive fusion proteins (NSF) and small, soluble attachment proteins (SNAP).
Because tissue distribution and VAMPs are more numerous and widely distributed than initially recognized, research on their differential expression and subcellular localization may turn out to be one of the most fruitful areas for the control or amelioration of diseases and disease symptoms.
Cellubrevins are associated with particular cell types, participate in both intracellular and extracellular pathways, and appear to vary in their abundance and specificity. Elucidation of the interactions of the novel cellubrevins (and associated VAMPs) with docking proteins such as syntaxin and SNAPs of the plasmalemma or the core fusion proteins such as NSF and the synaptotagmins (Bark I C and Wilson M C (1994) Proc Natl Acad Sci 91:4621-4624) provide means for the regulation of vesicle trafficking in normal as well as acute and chronic disease situations.
DISCLOSURE OF THE INVENTION
The subject invention provides nucleotide sequences which uniquely encode novel human cellubrevins. The cDNAs, disclosed herein, are designated: 1) cb-1 (SEQ ID NO:1) which was found within Incyte Clone No. 80184 and encodes a polypeptide designated CB-1 (SEQ ID NO:2); 2) cb-2 (SEQ ID NO:3) which was found within Incyte Clone No. 122826 and encodes a polypeptide designated CB-2 (SEQ ID NO:4), 3) cb-3 (SEQ ID NO:5) which was identified and extended using Incyte Clone No. 311537 and encodes a polypeptide designated CB-3 (SEQ ID NO:6); and 4) cb-4 (SEQ ID NO:7) which was found within Incyte Clone No. 674 719 and encodes a polypeptide designated CB-4 (SEQ ID NO:8).
The invention also comprises using these CBs or their variants to intercede in conditions involving physiologic or pathologic compromise which include the steps of testing a sample or an extract with cb nucleic acids, fragments or oligomers thereof. Aspects of the invention include the antisense of the nucleic acid sequences; cloning or expression vectors containing the nucleic acid sequences; host cells or organisms transformed with these expression vectors; a method for the production and recovery of purified CBs from host cells; and purified proteins which may be used to generate antibodies for diagnosis or therapy of activated or inflamed cells and/or tissues.
REFERENCES:
patent: 5650280 (1997-07-01), Stuart et al.
McMahon et al., “Cellubrevin is a ubiquitous tetanus-toxin substrate homologous to a putative synaptic vesicle fusion protein”,Nature, 364:346-349 (1993).
Arora, Naveen, “Cytotoxic Effects of a Chirmeric Protein Consisting of Tetanus Toxin Light Chain and Anthrax Toxin Lethal Factor in Non-neuronal Cells”,J. Biol. Chem., 269:26165-26171 (1994).
Bark et al., “Regulated vesicular fusion in neurons: Snapping together the details”,Proc. Natl. Acad. Sci. USA, 91:4621-4624 (1994).
Link et al., “Cleavage of Cellubrevin by Tetanus Toxin Does Not Affect Fusion of Early Endosomes”,J. Biol. Chem.,268-18423-18426 (1993).
Maniatis et al.,Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Press, Plainview, NY (1989).
Ausubel FM et al.,Current Protocols in Molecular Biology, John Wiley & Sons, New York, NY (1989).
Hohlfeld P. et al., “Prenatal diagnosis of congential toxoplasmosis with a polymerase chain reaction test on amniotic fluid”,New England Journal of Medicine, 331:695-699 (1994).
Mulder J. et al., “Rapid and simple PCR assay for quantitation of human immunodeficiency virus type 1 RNA in plasma:Application to acute retroviral infection”,J. Clin. Microbiol.31:292-300 (1994).
Ralston et al., “Expression of the Synaptic Vesicle Proteins VAMPs/Synaptobrevins 1 and 2 in Non-neural Tissues”,J. Biol. Chem.,269:15403-15406 (1994).
Archer et al., “Structures and chromosomal localizations of two human genes encoding synaptobrevins 1 and 2”,J. Biol. Chem.,vol. 265, No. 28, pp 17267-17273, (1990).
McMahon et al., “Cellubrevin: a substrate for tetanus toxin found on a constitutive vesicle recycling pathway is a homologue of synaptobrevin”, Soc. Neurosci. Abst., vol. 19, No. 1-3, p.900, (1993).
Sengupta et al., “Identification of novel isoforms of cellubrevin in rat pancreatic acinar cells”,Mol. Biol. Cell.,vol. 5, p.116A (1994).
Galli et al., “Cellubrevin is not restricted to transferrin receptor-containing vesicles in PC12 cells and colocalizes with synaptobrevin I and II”,Mol. Biol. Cell.,vol. 5, p. 325A (1994).
Galli et al., “Tetanus toxin-mediated cleavage of cellubrevin impairs exocytosis of transferrin receptor-contianing vesicles in CHO cells”,J. Cell. Biol.,vol. 125, No. 5, pp 1015-1024 (1994).
Jacobsson et al., “Identification of synaptic proteins and their isoform mRNAs in compartments of pancreatic endocrine cells”,Proc. Natl Acad. Sci.,91:12487-12491 (1994).
Sudhof et al., “A synaptic vesicle membrane protein is conserved from mammals to Drosophila”, Neuron, 2:1475-1481 (1989).
Baumert et al., “Synaptobrevin: an integral membrane protein of 18 000 daltons present in small synaptic vesicles of rat brain”,Embo. J.8(2): 379-384 (1989).
Richter, K. et al., “Gene expression in the embroyonic nervous system of Xenopus laevis”,Proc. Natl. Acad. Sci. USA85: 8086-90 (1988) (Accession GI 388483).
Hawkins Phillip R.
Murry Lynn E.
Seilhamer Jeffrey J.
Stuart Susan G.
Incyte Genomics, Corp.
Incyte Genomics, Corp.
Souaya Jehanne
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