ISG15 [untagged]


Catalogue Number
60-0008-500
Product Size
500 µg
Price £
£175
Accession Number
NP_005092
Residues Expressed
1-157
Certificate of Analysis Size
500 µg
Species
human
Source
E.coli
Quantity
500 µg
Storage
-70°C
Concentration
1 mg/ml
Formulation
50 mM HEPES pH 7.5, 150 mM sodium chloride, 2 mM dithiothreitol, 10% glycerol
Molecular Weight
~17.59 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_005092. For full protein sequence information download the Certificate of Analysis pdf.
QA; Protein Identification
Confirmed by mass spectrometry.
QA Activity

E1 Thioester ISG15 Loading Assay: The activity of ISG15 was validated by loading ISG15 onto the active cysteine of His-UBA7. Incubation of the His-UBA7 enzyme in the presence of ISG15 and ATP at 30°C was compared at two time points, T0 and T10 minutes.  Sensitivity of the ISG15/His-UBA7 thioester bond to the reducing agent DTT was confirmed.


Background

The enzymes of the ISGylation pathway play a pivotal role in the innate immune response. Three classes of enzymes are involved in the process of ISGylation; the Interferon-stimulated 15 kDa protein (ISG15) activating enzyme UBA7 (E1), the ISG15 conjugating enzyme UBE2L6 (E2) and the ubiquitin like modifier ISG15. ISG15 is a member of the ubiquitin like modifiers and the human gene was first described by Zhao et al. (2004). ISG15 functions in various biological pathways from pregnancy to innate immune responses (Ritchie and Zhang. 2004; Zhao et al., 2004). Secretion of ISG15 from monocytes in response to type I Interferons has been shown to cause natural killer (NK) cell proliferation and an increase in non-(major histocompatibility complex) MHC cytotoxicity (Meraro et al., 2002). ISG15 has also been shown to function intracellularly, being covalently conjugated to other proteins by UBA7, UBE2L6 and E3 ligases via a multistep process analogous to ubiquitylation (Loeb and Haas. 1992; Zhao et al., 2005). Ubiquitin-Specific Protease, 43kDa (UBP43) also known as Ubiquitin-Specific Protease, 18 (USP18) has been shown to specifically remove ISG15 from modified proteins (Malakhov et al., 2002).


References

Loeb KR, Haas AL (1992) The interferon-inducible 15-kDa ubiquitin homolog conjugates to intracellular proteins. J Biol Chem 267, 7806-13.

Malakhov MP, Malakhova OA, Kim KI, Ritchie KJ, Zhang DE (2002) UBP43 (USP18) specifically removes ISG15 from conjugated proteins. J Biol Chem 277, 9976-81.

Meraro D, Gleit-Kielmanowicz M, Hauser H, Levi BZ (2002) IFN-stimulated gene 15 is synergistically activated through interactions between the myelocyte/lymphocyte-specific transcription factors, PU.1, IFN regulatory factor-8/IFN consensus sequence binding protein, and IFN regulatory factor-4: characterization of a new subtype of IFN-stimulated response element. J Immunol 168, 6224-31.

Ritchie KJ, Zhang DE (2004) ISG15: the immunological kin of ubiquitin. Semin Cell Dev Biol 15, 237-46.

Zhao C, Beaudenon SL, Kelley ML, Waddell MB, Yuan W, Schulman BA, Huibregtse JM, Krug RM (2004) The UbcH8 ubiquitin E2 enzyme is also the E2 enzyme for ISG15, an IFN-alpha/beta-induced ubiquitin-like protein. Proc Natl Acad Sci USA 101, 7578-82.

Zhao C, Denison C, Huibregtse JM, Gygi S, Krug RM (2005) Human ISG15 conjugation targets both IFN-induced and constitutively expressed proteins functioning in diverse cellular pathways. Proc Natl Acad Sci USA 102, 10200-5.