Cloning, expression and purification of the α-carbonic anhydrase from the mantle of the Mediterranean mussel, Mytilus galloprovincialis.
J Enzyme Inhib Med Chem
; 32(1): 1029-1035, 2017 Dec.
Article
em En
| MEDLINE
| ID: mdl-28741386
ABSTRACT
We cloned, expressed, purified, and determined the kinetic constants of the recombinant α-carbonic anhydrase (rec-MgaCA) identified in the mantle tissue of the bivalve Mediterranean mussel, Mytilus galloprovincialis. In metazoans, the α-CA family is largely represented and plays a pivotal role in the deposition of calcium carbonate biominerals. Our results demonstrated that rec-MgaCA was a monomer with an apparent molecular weight of about 32 kDa. Moreover, the determined kinetic parameters for the CO2 hydration reaction were kcat = 4.2 × 105 s-1 and kcat/Km of 3.5 × 107 M-1 ×s-1. Curiously, the rec-MgaCA showed a very similar kinetic and acetazolamide inhibition features when compared to those of the native enzyme (MgaCA), which has a molecular weight of 50 kDa. Analysing the SDS-PAGE, the protonography, and the kinetic analysis performed on the native and recombinant enzyme, we hypothesised that probably the native MgaCA is a multidomain protein with a single CA domain at the N-terminus of the protein. This hypothesis is corroborated by the existence in mollusks of multidomain proteins with a hydratase activity. Among these proteins, nacrein is an example of α-CA multidomain proteins characterised by a single CA domain at the N-terminus part of the entire protein.
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Texto completo:
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Anidrases Carbônicas
/
Mytilus
Limite:
Animals
Idioma:
En
Ano de publicação:
2017
Tipo de documento:
Article