Carboxypeptidase A1/CPA1 Protein, Mouse (HEK293, His)

Carboxypeptidase A1 (CPA1) is an enzyme responsible for catalyzing the release of a C-terminal amino acid; however, it exhibits limited or no activity with residues such as -Asp, -Glu, -Arg, -Lys, or -Pro. This specificity in substrate recognition underscores its role as a carboxypeptidase with selectivity towards certain amino acid sequences at the C-terminus of peptides. Carboxypeptidase A1/CPA1 Protein, Mouse (HEK293, His) is the recombinant mouse-derived Carboxypeptidase A1/CPA1 protein, expressed by HEK293 , with C-His labeled tag.

Carboxypeptidase A1 (CPA1) is an enzyme that plays a crucial role in catalyzing the release of C-terminal amino acids from peptides. However, its activity is limited or absent for peptides with C-terminal residues such as -Asp, -Glu, -Arg, -Lys, or -Pro. This substrate specificity indicates that CPA1 exhibits selectivity in its cleavage preferences, preferring certain amino acid residues at the C-terminus of peptides. The enzyme's action in releasing C-terminal amino acids contributes to various physiological processes, such as the digestion of proteins in the digestive system, highlighting its importance in the regulation of peptide and protein metabolism.

Measured by its ability to cleave the colorimetric peptide substrate Ac-Phe-Thiaphe-OH in the presence of 5,5’Dithio-bis (2-nitrobenzoic acid) (DTNB). The specific activity is 5420.437 pmol/min/µg, as measured under the described conditions. (Activation description: The proenzyme needs to be activated by Trypsin for an activated form.)

Materials
Assay buffer: 50 mM Tris, 0.15 M NaCl, 10 mM CaCl₂, 0.05% Brij-35, pH 7.5 (TCNB)
Carboxypeptidase A1/CPA1 Protein, Mouse (HEK293, His) (HY-P72868)
Trypsin
Substrate: Ac-Phe-Thiaphe-OH
DTNB (HY-15915)

Procedure
1. Dilute Mouse CPA1 to 100 µg/mL using a detection buffer containing 1.0 µg/mL Trypsin.
2. Incubate at room temperature for 60 minutes. Dilute Mouse CPA1 to 0.2 µg/mL in the assay buffer.
3. Mix equal volumes of 10 mM substrate and 10 mM DTNB. Then dilute the mixture to 200 µM in the assay buffer.
4. Add 50 µL of 0.2 µg/mL Mouse CPA1 to a transparent microplate. Add 50 µL of assay buffer to the blank.
5. Add 50 µL of the 200 µM substrate mixture to the wells to start the reaction.
6. Read at 405 nm in kinetic mode for 5 minutes.
7. Calculate specific activity:

Mouse

HEK293

C-His

Q7TPZ8 (N17-Y419)

109697  [NCBI]

Full Length of Mature Protein

NENFVGHQVLRISATDEAQVQKVRELEELEHLKLDFWRDPARAGLPIDVRVPFPTIQSVKAFLEYHDISYEIMIEDVQSLLDEEKQQMSAFQARALSTDAFNYATYHTLDEIYEFMDLLVTEHPQLVSKIQIGSTFEGRPINVLKFSTGGTNRPAIWIDTGIHSREWVTQASGVWFAKKITKDYGQEPTLTAILDNMDIFLEIVTNPDGFVYTHKTNRMWRKTRSHTEGSLCVGVDPNRNWDAAFGMPGASSNPCSETYRGKFPNSEVEVKSIVDFVTSHGNIKAFISIHSYSQLLLYPYGYTSEPAPDKEELDQLAKSAVTALTSLHGTKFKYGSIIDTIYQASGSTIDWTYSQGIKYSFTFELRDTGLRGFLLPASQIIPTAEETWLALLTIMDHTVKHPY

Approximately 42-49 kDa

• 
  Greater than 95% as determined by reducing SDS-PAGE.

Lyophilized powder

Lyophilized from a 0.22 μm filtered solution of 20 mM Tris, 150 mM NaCl, pH 7.5 or 25 mM Tris-HCl, 150 mM NaCl, pH 7.5.
Note: For SPR assay, please replace the buffer. Primary amine components (e.g., Tris, imidazole) can affect protein-coupled chips.

<1 EU/μg, determined by LAL method.

It is not recommended to reconstitute to a concentration less than 100 μg/mL in ddH₂O.

Stored at -20°C for 2 years from date of receipt. After reconstitution, it is stable at 4°C for 1 week or -20°C for longer (with carrier protein). It is recommended to freeze aliquots at -20°C or -80°C for extended storage.

Room temperature in continental US; may vary elsewhere.