ADAMTS-4, Recombinant his tag

Description
Catalogue Number	CC1028
Brand Family	Chemicon®
Trade Name	Chemicon
Description	ADAMTS-4, recombinant, His Tagged
Overview	Molecular form: Recombinant human ADAMTS4 delta580-837 is produced with the baculovirus expression system and purified from insect cell culture supernatants. The protein consists of amino acids F213 to A579 of full-length ADAMTS4 and a C-terminal His6tag. The calculated Mr is 40,366 Da. Inhibitors: ADAMTS4 is inhibited by tissue inhibitors of matrix metalloproteinases 3 (TIMP 3) and to a lesser extend by other tissue inhibitors. Enzyme activity is also suppressed by chelators of divalent captions as EDTA and by synthetic metalloproteinase inhibitors. Applications: Recombinant ADAMTS4 is used to study the degradation of extracellular matrix proteoglycans, to screen for inhibitors of proteoglycan hydrolysis and to characterize inhibitor actions. The enzyme can also serve as standard in enzymatic and immunochemical assays.
Alternate Names	Aggrecanase 1
Background Information	ADAMTS (a disintegrin and metalloproteinase with thrombospondin motif) is a novel family of extrcellular proteinases (Tang, 2001). Presently nine family members have been identified in mammals (Tang, 2001). ADAMTS4 was first purified from IL-1-stimulated bovine nasal cartilage conditioned media and human ADAMTS4-cDNA was cloned from a human heart cDNA library (Tortorella, 1999). Mature ADAMTS4 consists of a prodomain which confers latency to the proenzyme, a catalytic domain, a disintegrin domain and a C-terminal sequence with a thrombospondin type-1 motif. The prodomain is most likely cleaved off by a furin-type enzyme before active ADAMTS4 is released from cells. Active ADAMTS4 consists of 625 amino acids (F213 - K837) with a calculated Mr of 67 943 (Tortorella, 1999). ADAMTS4 hydrolyzes aggrecan, the major proteoglycan of articular cartilage (Tortorella, 1999). As aggrecan is also digested by 2 other members of the ADAMTS family, ADAMTS1 (Kuno, 2000) and ADAMTS5 (Abbaszade, 1999), aggrecan degradation products found in normal and rheumatoid and osteoarthritic joint cartilage (Lark, 1997) may arise from action of either of the 3 proteinases. Isolated ADAMTS4 hydrolyzes aggrecan at 5 different sites (Tortorella, 2000a). Four cleavage sites are located in the chondroitin sulfate-rich region between globular domains G2 and G3, while one site is placed in the rod-shaped polypeptide between globular domains G1 and G2. The thrombospondin motif in ADAMTS4 appears to be critical for aggrecan substrate recognition and cleavage (Tortorella, 2000b). ADAMTS4 hydrolyzes also other lecticans as brevican (Nakamura, 2000) and versican (Sandy, 2001). ADAMTS4 is inhibited by TIMP 3 (Hashimoto, 2001) and by the N-terminal domain of TIMP 3 (Kashigawa, 2001) with Ki-values in the nanomolar range. Inhibition by TIMP 1,2, and 4 is much weaker (Hashimoto, 2001).

Description

Catalogue Number

CC1028

Brand Family

Chemicon®

Trade Name

Chemicon

Description

ADAMTS-4, recombinant, His Tagged

Overview

Molecular form: Recombinant human ADAMTS4 delta580-837 is produced with the baculovirus expression system and purified from insect cell culture supernatants. The protein consists of amino acids F213 to A579 of full-length ADAMTS4 and a C-terminal His6tag. The calculated Mr is 40,366 Da.

Inhibitors: ADAMTS4 is inhibited by tissue inhibitors of matrix metalloproteinases 3 (TIMP 3) and to a lesser extend by other tissue inhibitors. Enzyme activity is also suppressed by chelators of divalent captions as EDTA and by synthetic metalloproteinase inhibitors.

Applications: Recombinant ADAMTS4 is used to study the degradation of extracellular matrix proteoglycans, to screen for inhibitors of proteoglycan hydrolysis and to characterize inhibitor actions. The enzyme can also serve as standard in enzymatic and immunochemical assays.

Alternate Names

Aggrecanase 1

Background Information

ADAMTS (a disintegrin and metalloproteinase with thrombospondin motif) is a novel family of extrcellular proteinases (Tang, 2001). Presently nine family members have been identified in mammals (Tang, 2001). ADAMTS4 was first purified from IL-1-stimulated bovine nasal cartilage conditioned media and human ADAMTS4-cDNA was cloned from a human heart cDNA library (Tortorella, 1999). Mature ADAMTS4 consists of a prodomain which confers latency to the proenzyme, a catalytic domain, a disintegrin domain and a C-terminal sequence with a thrombospondin type-1 motif. The prodomain is most likely cleaved off by a furin-type enzyme before active ADAMTS4 is released from cells. Active ADAMTS4 consists of 625 amino acids (F213 - K837) with a calculated Mr of 67 943 (Tortorella, 1999).

ADAMTS4 hydrolyzes aggrecan, the major proteoglycan of articular cartilage (Tortorella, 1999). As aggrecan is also digested by 2 other members of the ADAMTS family, ADAMTS1 (Kuno, 2000) and ADAMTS5 (Abbaszade, 1999), aggrecan degradation products found in normal and rheumatoid and osteoarthritic joint cartilage (Lark, 1997) may arise from action of either of the 3 proteinases. Isolated ADAMTS4 hydrolyzes aggrecan at 5 different sites (Tortorella, 2000a). Four cleavage sites are located in the chondroitin sulfate-rich region between globular domains G2 and G3, while one site is placed in the rod-shaped polypeptide between globular domains G1 and G2. The thrombospondin motif in ADAMTS4 appears to be critical for aggrecan substrate recognition and cleavage (Tortorella, 2000b). ADAMTS4 hydrolyzes also other lecticans as brevican (Nakamura, 2000) and versican (Sandy, 2001).

ADAMTS4 is inhibited by TIMP 3 (Hashimoto, 2001) and by the N-terminal domain of TIMP 3 (Kashigawa, 2001) with Ki-values in the nanomolar range. Inhibition by TIMP 1,2, and 4 is much weaker (Hashimoto, 2001).

Product Information
Presentation	ADAMTS4 is solublized in 50mM Tris-HCI, pH 7.5, 150 mM NaCl, 5mM CaCl2, 0.05% Brij-35.

Product Information

Presentation

ADAMTS4 is solublized in 50mM Tris-HCI, pH 7.5, 150 mM NaCl, 5mM CaCl2, 0.05% Brij-35.

Applications
Application	This His Tagged Recombinant ADAMTS4 is used to study the degradation of extracellular matrix proteoglycans, to screen for inhibitors of proteoglycan hydrolysis and to characterize inhibitor actions.
Key Applications	Affects Function

Applications

Application

This His Tagged Recombinant ADAMTS4 is used to study the degradation of extracellular matrix proteoglycans, to screen for inhibitors of proteoglycan hydrolysis and to characterize inhibitor actions.

Key Applications

Affects Function

Biological Information
Purity	Recombinant ADAMTS4 appears as a major protein of about 42,000 Da in SDS-PAGE (> 90% of total protein). Due to autoproteolytic activity minor bands of ADAMTS4 may be visible in the enzyme preparation.
Source	Recombinant human ADAMTS-4 δ580-837 is produced with the baculovirus expression system and purified from insect cell culture supernatant. The protein contains amino acids F213 ... A579 of full-length ADAMTS-4 and a C-terminal His6tag. Calculated MW is 40 kDa.
Specific Activity	Specific Activity: Aggrecanase activity of ADAMTS4 is determined with recombinant His-tagged aggrecan interglobular domain (Aggrecan-IGD1 from Chemicon). ADAMTS4 hydrolyzes the "aggrecanase" site within this domain (peptide bond E373 -A374 in human aggrecan). The recombinant substrate is incubated at a concentration of 0.1 μM with 1 nM ADAMTS4 in 50 mM Tris-HCI, pH 7.5, 150 mM NaCl, 5 mM CaCl2, 1 μM leupeptin, 1 μM pepstatin, 1mM Pefabloc, 0.05% Brij 35 for 15 mins at 37°C. Cleavage at the "aggrecanase" - site is estimated from the appearance of the C-terminus ARGSVIL. Using polyclonal neoepitope antibodies to the ARGS-terminus the fragment is fixed to a microplate and quantified with anti-His-tag antibody. Under the specified conditions, recombinant ADAMTS4 hydrolyzes 0.15 nM substrate/ min per nM truncated ADAMTS4. When related to mg enzyme the value is 3.7 nmoles hydrolyzed substrate/min per mg ADAMTS4.
Entrez Gene Number	NM_005099.4
Entrez Gene Summary	This gene encodes a member of the ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) protein family. Members of the family share several distinct protein modules, including a propeptide region, a metalloproteinase domain, a disintegrin-like domain, and a thrombospondin type 1 (TS) motif. Individual members of this family differ in the number of C-terminal TS motifs, and some have unique C-terminal domains. The enzyme encoded by this gene lacks a C-terminal TS motif. It is responsible for the degradation of aggrecan, a major proteoglycan of cartilage, and brevican, a brain-specific extracellular matrix protein. The cleavage of aggrecan and brevican suggests key roles of this enzyme in arthritic disease and in the central nervous system, potentially, in the progression of glioma.
Gene Symbol	ADAMTS4 ADAM-TS4 aggrecanase-1 ADAMTS-2 ADMP-1 KIAA0688 Aggrecanase-1 ADAMTS-4 EC 3.4.24.82
UniProt Number	O75173
UniProt Summary	FUNCTION: SwissProt: O75173 # Cleaves aggrecan, a cartilage proteoglycan, and may be involved in its turnover. May play an important role in the destruction of aggrecan in arthritic diseases. Could also be a critical factor in the exacerbation of neurodegeneration in Alzheimer disease. Cleaves aggrecan at the '392-Glu- -Ala-393' site. COFACTOR: Binds 1 zinc ion per subunit (By similarity). SIZE: 837 amino acids; 90225 Da SUBCELLULAR LOCATION: Secreted, extracellular space, extracellular matrix (By similarity). TISSUE SPECIFICITY: Expressed in brain, lung and heart. Expressed at very low level in placenta and skeletal muscles. DOMAIN: SwissProt: O75173 The spacer domain and the TSP type-1 domains are important for a tight interaction with the extracellular matrix. & The conserved cysteine present in the cysteine-switch motif binds the catalytic zinc ion, thus inhibiting the enzyme. The dissociation of the cysteine from the zinc ion upon the activation-peptide release activates the enzyme. PTM: The precursor is cleaved by a furin endopeptidase. SIMILARITY: Contains 1 disintegrin domain. & Contains 1 peptidase M12B domain. & Contains 1 TSP type-1 domain.

Biological Information

Purity

Recombinant ADAMTS4 appears as a major protein of about 42,000 Da in SDS-PAGE (> 90% of total protein). Due to autoproteolytic activity minor bands of ADAMTS4 may be visible in the enzyme preparation.

Source

Recombinant human ADAMTS-4 δ580-837 is produced with the baculovirus expression system and purified from insect cell culture supernatant. The protein contains amino acids F213 ... A579 of full-length ADAMTS-4 and a C-terminal His6tag. Calculated MW is 40 kDa.

Specific Activity

Specific Activity: Aggrecanase activity of ADAMTS4 is determined with recombinant His-tagged aggrecan interglobular domain (Aggrecan-IGD1 from Chemicon). ADAMTS4 hydrolyzes the "aggrecanase" site within this domain (peptide bond E373 -A374 in human aggrecan). The recombinant substrate is incubated at a concentration of 0.1 μM with 1 nM ADAMTS4 in 50 mM Tris-HCI, pH 7.5, 150 mM NaCl, 5 mM CaCl2, 1 μM leupeptin, 1 μM pepstatin, 1mM Pefabloc, 0.05% Brij 35 for 15 mins at 37°C. Cleavage at the "aggrecanase" - site is estimated from the appearance of the C-terminus ARGSVIL. Using polyclonal neoepitope antibodies to the ARGS-terminus the fragment is fixed to a microplate and quantified with anti-His-tag antibody. Under the specified conditions, recombinant ADAMTS4 hydrolyzes 0.15 nM substrate/ min per nM truncated ADAMTS4. When related to mg enzyme the value is 3.7 nmoles hydrolyzed substrate/min per mg ADAMTS4.

Entrez Gene Number

NM_005099.4

Entrez Gene Summary

This gene encodes a member of the ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) protein family. Members of the family share several distinct protein modules, including a propeptide region, a metalloproteinase domain, a disintegrin-like domain, and a thrombospondin type 1 (TS) motif. Individual members of this family differ in the number of C-terminal TS motifs, and some have unique C-terminal domains. The enzyme encoded by this gene lacks a C-terminal TS motif. It is responsible for the degradation of aggrecan, a major proteoglycan of cartilage, and brevican, a brain-specific extracellular matrix protein. The cleavage of aggrecan and brevican suggests key roles of this enzyme in arthritic disease and in the central nervous system, potentially, in the progression of glioma.

Gene Symbol

ADAMTS4
ADAM-TS4
aggrecanase-1
ADAMTS-2
ADMP-1
KIAA0688
Aggrecanase-1
ADAMTS-4
EC 3.4.24.82

UniProt Number

O75173

UniProt Summary

FUNCTION: SwissProt: O75173 # Cleaves aggrecan, a cartilage proteoglycan, and may be involved in its turnover. May play an important role in the destruction of aggrecan in arthritic diseases. Could also be a critical factor in the exacerbation of neurodegeneration in Alzheimer disease. Cleaves aggrecan at the '392-Glu- -Ala-393' site.
COFACTOR: Binds 1 zinc ion per subunit (By similarity).
SIZE: 837 amino acids; 90225 Da
SUBCELLULAR LOCATION: Secreted, extracellular space, extracellular matrix (By similarity).
TISSUE SPECIFICITY: Expressed in brain, lung and heart. Expressed at very low level in placenta and skeletal muscles.
DOMAIN: SwissProt: O75173 The spacer domain and the TSP type-1 domains are important for a tight interaction with the extracellular matrix. & The conserved cysteine present in the cysteine-switch motif binds the catalytic zinc ion, thus inhibiting the enzyme. The dissociation of the cysteine from the zinc ion upon the activation-peptide release activates the enzyme.
PTM: The precursor is cleaved by a furin endopeptidase.
SIMILARITY: Contains 1 disintegrin domain. & Contains 1 peptidase M12B domain. & Contains 1 TSP type-1 domain.

Product Usage Statements
Usage Statement	Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Product Usage Statements

Usage Statement

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Storage and Shipping Information
Storage Conditions	Maintain protein at -70°C for one year from date of receipt. The enzyme can be kept at -20°C for several weeks, and on ice for several days. Repeated freezing and thawing should be avoided.

Storage and Shipping Information

Storage Conditions

Maintain protein at -70°C for one year from date of receipt. The enzyme can be kept at -20°C for several weeks, and on ice for several days. Repeated freezing and thawing should be avoided.

Packaging Information
Material Size	5 µg
Material Package	in 25 μL

Packaging Information

Material Size

5 µg

Material Package

in 25 μL

实验耗材

分子生物学试剂

蛋白与免疫学

生化试剂

细胞及检测试剂

色谱及层析

仪器设备

实验服务

办公用品

ADAMTS-4, Recombinant his tag

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