Production and characterization of a novel aminopeptidase A from Lactococcus lactis / 生物工程学报

Aminopeptidase A (Pep A) is a metal-dependent enzyme that specifically hydrolyze peptides with the N-terminal amino acids glutamic acid (Glu) and aspartic acid (Asp). A possible application of PepA is the hydrolysis of Glu/Asp-rich food proteins such as wheat gluten and casein, increasing the flavor and solubility of food protein. In the present study, the gene encoding a Pep A from Lactococcus lactis ssp. lactis IL1403 was synthesized and introduced into Pichia pastoris GS115 (His4). Lc-Pep A was successfully expressed and secreted to the culture medium, followed by identification and purification to homogeneity. Characteristics study demonstrated that Lc-Pep A could specifically hydrolyze the substrates Glu-pNA and Asp-pNA with similar catalytic activity, and this was further confirmed by the kinetics parameters measured. Additionally, Lc-Pep A showed a broad thermostability and pH stability with an optimum temperature of 60 ℃ and an optimum pH of 8.0. The enzyme activity of Lc-Pep A was activated by metal ions Co2+, Mn2+, and Zn2+ but was strongly inhibited by Ni2+and Cu2+. The routine proteinase inhibitor had no effect on the activity of Lc-Pep A. However, Lc-Pep A was strongly inhibited by the metallopeptidase inhibitor, EDTA, and disulfide bond-reducing agents. The study may facilitate production and application of Lc-Pep A.

Effects of Angiotensin III in Rabbit Corpus Cavernosum Smooth Muscle Contraction: Comparing with Angiotensin I and Angiotensin II / 대한비뇨기과학회지

PURPOSE: Angiotensin(ANG) II regulates tone of penile smooth muscle for erection. ANG III is a product converted from ANG II by aminopeptidase A. The effects of ANG III have not been clarified in the penile corpus cavernosum. The purpose of the present experiment was to determine whether the ANG III has regulatory function in the control of rabbit corpus cavernosum smooth muscle tone. MATERIALS AND METHODS: A strip of rabbit corpus cavernosum was mounted in an organ chamber to measure the isometric tension. We compared the effects of ANG III(10-7M to 10-5M), ANG II(10-8M to 10-6M) and ANG I(10-7M to 10-5M) on the contractility of the corpus cavernosum smooth muscle. RESULTS: ANG III, ANG II, and ANG I contracted corpus cavernosum smooth muscle strips dose-dependently. The contraction of smooth muscle induced by ANG III was 10 fold less by ANG II. Contractile response to ANG III was not attenuated by captopril(angiotensin converting enzyme inhibitor). Contractile response to ANG III was significantly inhibited by Dup 753 of 10-7M(type 1 specific ANG II receptor inhibitor) but not inhibited by PD 123,319 of 10-6M(type 2 specific ANG II inhibitor). CONCLUSIONS: The present results suggest that ANG III is involved in the regulation of corpus cavernosum smooth muscle tone, and contractile effect to ANG III produced via activation of type 1 ANG II (AT1) receptor. The rank order of potency of contraction was as follows, ANG II>ANG IIIANG I.