Artigo de peri??dico
Structural characterization and enzymatic activity of the recombinant Ala959 to Ser1066 region of human ace
Autor
ELIASA, CAROLINE C.
PEREIRA, LARISSA M.
ARAGAO, DANIELLE S.
CASARINI, DULCE E.
AFFONSO, REGINA
Resumen
Angiotensin-converting enzyme catalyzes the conversion of angiotensin I to the vasoconstrictor angiotensin II and the hydrolysis
of bradykinin (BK). Human somatic angiotensin-converting enzyme has two homologous domains (N and C) that share 60%
identity. Although these two regions have high homology, the catalytic site of the C-domain exhibits three-fold greater activity
than the N-domain in the hydrolysis of angiotensin I in vivo. The present study aimed to obtain the Ala959 to Ser1066 catalytic region
of the C-domain of angiotensin-converting enzyme in a structural conformation that resembles its native form. We amplified the
324-bp sequence corresponding to the catalytic site of C-domain of angiotensin-converting enzyme and cloned this sequence
into a pET28 vector. The catalytic site of C-domain of angiotensin-converting enzyme peptide was expressed in a bacterial system,
and its purification was performed in one step using a His-tag affinity column. Structural analysis by circular dichroism and
fluorescence confirmed that the purified protein is correctly folded, and catalytic site of C-domain of angiotensin-converting
enzyme possesses enzymatic activity and is inhibited by lisinopril. This peptide can be used to test new inhibitors and C-domain
of angiotensin-converting enzyme substrates because this peptide is easy to produce and this has proven efficient link with these
molecules.