Intracellular proteolytic processing of proopiomelanocortin in heterologous COS-1 cells by the yeast KEX2 endoprotease

1990 ◽  
Vol 68 (3) ◽  
pp. 635-640 ◽  
Author(s):  
L. Zollinger ◽  
C. Racine ◽  
P. Crine ◽  
G. Boileau ◽  
D. Germain ◽  
...  
Keyword(s):  
High Pressure ◽  
Liquid Chromatography ◽  
High Pressure Liquid Chromatography ◽  
Reversed Phase ◽  
Proteolytic Processing ◽  
Amino Terminal ◽  
Cell Extracts ◽  
Gel Permeation

We have transiently expressed the yeast KEX2 gene together with the proopiomelanocortin (POMC) cDNA in COS-1 cells. Characterization of the POMC-related immunoreactive peptides by gel permeation and reversed-phase high pressure liquid chromatography showed that the KEX2 enzyme was active and capable of carrying out cleavage of POMC to release the authentic maturation product β-endorphin(1–31). Peptides resembling β-lipotropin, the amino terminal glycopeptide, and ACTH(1–39) were also detected as major products in the cell extracts. Our results indicate that the KEX2 enzyme can proteolytically release from POMC a set of peptides similar to that normally found in interior pituitary.Key words: prohormone proteolytic processing, KEX2 expression, COS-1 cells.

scholarly journals Ekstraksi, Karakterisasi dan Uji Aktivitas Antioksidan Astaxanthin dari Produk Fermentasi Udang (Cincalok)

2021 ◽  
Vol 24 (3) ◽  
pp. 311-322
Author(s):  
Mauludia Mauludia ◽  
Thamrin Usman ◽  
Winda Rahmalia ◽  
Dwi Imam Prayitno ◽  
Siti Nani Nurbaeti
Keyword(s):  
Antioxidant Activity ◽  
High Pressure ◽  
Liquid Chromatography ◽  
High Pressure Liquid Chromatography ◽  
Aquatic Organisms ◽  
Dpph Method ◽  
Wet Weight ◽  
Tlc Analysis ◽  
Layer Chromatography

Shrimp is one of the aquatic organisms that contain several active compounds, including astaxanthin. Cincalok is one of the fermented shrimp products containing astaxanthin. This study aims to determine the characteristics of astaxanthin extract from cincalok and its antioxidant activity. Extraction of astaxanthin from cincalok was carried out using the reflux method with acetone : cyclohexane (20:80 v/v) as a solvent. The identification and characterization of astaxanthin was carried out using thin-layer chromatography (TLC), UV-Vis spectrophotometry, and High-Pressure Liquid Chromatography (HPLC). Meanwhile, the antioxidant activity test was carried out using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) method in one serial concentration (5; 15; 25 ppm). The results of TLC analysis showed that astaxanthin in cincalok extract has Rf value (0.32). The analysis using a UV-Vis spectrophotometer produced a spectrum with a maximum wavelength of 477 nm, which corresponds to the maximum wavelength of standard astaxanthin. The yield of astaxanthin extract from cincalok in this study was 1.47 mg/100 g wet weight. The chromatogram from the results of UHPLC analysis showed that the retention time of cincalok astaxanthin extract was 6.27 minutes with a purity of 18.03%. The antioxidant activity of cincalok astaxanthin extract was 568.32 ppm. Udang merupakan salah satu organisme air yang mengandung banyak senyawa aktif, termasuk astaxanthin. Cincalok merupakan salah satu produk hasil fermentasi udang yang mengandung astaxanthin. Penelitian ini bertujuan untuk mengetahui karakteristik ekstrak astaxanthin dari cincalok dan aktivitas antioksidannya. Ekstraksi astaxanthin dari cincalok menggunakan metode refluks dengan pelarut aseton:sikloheksan (20:80 v/v). Identifikasi dan karakterisasi astaxanthin dilakukan dengan menggunakan kromatografi lapis tipis (KLT), spektrofotometri UV-Vis, dan High Pressure Liquid Chromatography (HPLC). Sedangkan uji aktivitas antioksidan dilakukan menggunakan metode 1,1-difenil-2-pikrilhidrazil (DPPH) dengan memvariasikan konsentrasi larutan uji, yaitu 5; 15; 25 ppm. Hasil dari penelitian ini melaporkan astaxanthin pada ekstrak cincalok menunjukkan nilai Rf 0,32 pada kromatografi lapis tipis (KLT). Hasil analisis menggunakan spektrofotometer UV-Vis menghasilkan spektra dengan panjang gelombang maksimum 477 nm, yang sesuai dengan panjang gelombang maksimum astaxanthin standar. Randemen ekstrak astaxanthin dari cincalok pada penelitian ini adalah 1,47 mg/100 g berat basah. Kromatogram dari hasil analisis UHPLC menunjukkan waktu retensi ekstrak astaxanthin cincalok yaitu selama 6,27 menit dengan kemurnian sebesar 18,03%. Aktivitas antioksidan dari ekstrak astaxanthin cincalok diperoleh sebesar 568,32 ppm.  

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