HSPA8/HSC70 Protein, Human (His, Solution)

HSPA8/HSC70 is a molecular chaperone that protects the proteome and aids in peptide folding, transport, chaperone-mediated autophagy, and protein complex regulation. It is central to quality control, ensuring correct folding and targeting misfolded proteins for degradation through ATP-dependent cycles. HSPA8/HSC70 Protein, Human (His, Solution) is the recombinant human-derived HSPA8/HSC70 protein, expressed by E. coli , with N-His labeled tag.

HSPA8/HSC70, a molecular chaperone, is intricately involved in diverse cellular processes, including proteome protection from stress, facilitation of polypeptide folding and transport, chaperone-mediated autophagy, activation of misfolded protein proteolysis, and modulation of protein complex formation and dissociation. Central to the protein quality control system, it ensures correct protein folding, refolding of misfolded proteins, and regulates protein targeting for subsequent degradation. This function is orchestrated through cycles of ATP binding, ATP hydrolysis, and ADP release, facilitated by co-chaperones. The nucleotide-bound state of HSP70 regulates its affinity for polypeptides, with ATP-bound form having low substrate affinity, and a conformational change upon ATP hydrolysis increasing the affinity for substrates. Co-chaperones, including J-domain co-chaperones (HSP40s), nucleotide exchange factors (NEFs) such as BAG1/2/3, and TPR domain chaperones like HOPX and STUB1, play specific roles in modulating HSP70 activity. Beyond its fundamental role in mitochondrial import, HSPA8/HSC70 also acts as a repressor of transcriptional activation, participates in the spliceosome assembly, and plays a role in selective protein degradation processes, including chaperone-mediated autophagy and ER-associated degradation. Additionally, it interacts with the VGF-derived peptide TLQP-21, indicating its involvement in diverse cellular pathways.

Hsp70 has ATPase activity at the time of manufacture of 1.514 µM phosphate liberated/h/µg protein in a 200 µL reaction at 37°C in the presence of 10 μL of 4mM ATP using a Malachite Green assay.

Materials
Test protein: HSPA8/HSC70 Protein, Human (His, Solution) (HY-P73915)
ATPase/GTPase Assay Kit
Adenosine 5′-triphosphate (ATP) disodium salt hydrate

Procedure
1. Take out the reagents from the ATPase/GTPase Assay Kit and equilibrate at room temperature for 30 minutes.
2. Prepare the standard phosphate solution at concentrations of 50, 30, 15, and 0 μM, and add 40 μL/well to the 96-well plate. The preparation details are as follows:

Human

E. coli

N-His

P11142-1 (M1-D646)

3312  [NCBI]

Full Length of Isoform-1

MSKGPAVGIDLGTTYSCVGVFQHGKVEIIANDQGNRTTPSYVAFTDTERLIGDAAKNQVAMNPTNTVFDAKRLIGRRFDDAVVQSDMKHWPFMVVNDAGRPKVQVEYKGETKSFYPEEVSSMVLTKMKEIAEAYLGKTVTNAVVTVPAYFNDSQRQATKDAGTIAGLNVLRIINEPTAAAIAYGLDKKVGAERNVLIFDLGGGTFDVSILTIEDGIFEVKSTAGDTHLGGEDFDNRMVNHFIAEFKRKHKKDISENKRAVRRLRTACERAKRTLSSSTQASIEIDSLYEGIDFYTSITRARFEELNADLFRGTLDPVEKALRDAKLDKSQIHDIVLVGGSTRIPKIQKLLQDFFNGKELNKSINPDEAVAYGAAVQAAILSGDKSENVQDLLLLDVTPLSLGIETAGGVMTVLIKRNTTIPTKQTQTFTTYSDNQPGVLIQVYEGERAMTKDNNLLGKFELTGIPPAPRGVPQIEVTFDIDANGILNVSAVDKSTGKENKITITNDKGRLSKEDIERMVQEAEKYKAEDEKQRDKVSSKNSLESYAFNMKATVEDEKLQGKINDEDKQKILDKCNEIINWLDKNQTAEKEEFEHQQKELEKVCNPIITKLYQSAGGMPGGMPGGFPGGGAPPSGGASSGPTIEEVD

Approximately 65-73 kDa, based on SDS-PAGE under reducing conditions.

• 
  ≥ 90%, as determined by reducing SDS-PAGE.

Solution

Supplied as a 0.22 μm filtered solution of PBS, 10% glycerol, pH 7.5.

<1 EU/μg, determined by LAL method.

N/A.

Stored at -80°C for 1 year from date of receipt. It is stable at -20°C for 3 months after opening. It is recommended to freeze aliquots at -80°C for extended storage. Avoid repeated freeze-thaw cycles.

Shipping with dry ice.