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PM · Lens · An Aspartic‑metalloprotease from an Endemic Plant Tuber (Burnatia enneandra micheli): Purification and Biochemical Characterization

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Article · Volume 14, Issue 57S · pp. S319–S326

An Aspartic‑metalloprotease from an Endemic Plant Tuber (Burnatia enneandra micheli): Purification and Biochemical Characterization

Ngangoum Eric Serge2,3, Mezajoug Kenfack Laurette Blandine2, Sanjit Kumar3, Tchiegang Clerge2, Mookambeswaran Vijayalakshmi3

  1. 1Department of Food Sciences and Nutrition, Bioprocess Laboratory, Institute University of Technology, University of Ngaoundere, Ngaoundéré, Cameroon, India.
  2. 2Centre for Bio‑Separation Technology, VIT University, Vellore, Tamil Nadu, India.

Abstract

Background: The objective of this work was to isolate, optimize, and characterize protease from Burnatia enneandra which is an endemic plant found abundantly in the Far‑Nord Region of Cameroon. The optimum condition to extract maximum quantity of protease from B. enneandra with respect to pH, the ratio (m/v), and agitation frequency was defined as 5.1%, 4%, and 100 rpm, respectively.Materials and Methods: The enzyme was purified using ammonium sulphate precipitation, double gel filtration chromatography sephadex G200 followed by sephadex G75 and the purified protease was further characterized. With an apparent molecular weight of 23 kDa on SDS-PAGE, the purified protease showed maximum activity at 5.1 and 40°C respectively for pH and temperature. Its activity was enhanced by metal ions such as Ca2+ and Ni2+, while Fe2+and Zn2+ showed significant inhibition.Results: B. enneandra protease activity was not affected by proteases inhibitors such as phenylmethylsulfonyl fluoride, aprotinin, and iodoacetamide but was strongly inhibited by Pepstatin A and ethylenediaminetetraacetic acid which allowed to classify this new protease as aspartic‑metalloproteases. Using casein as substrate, protease from B. enneandra had a maximum rate of reaction (Vmax) and Michaelis‑Menten constant (Km) of 64.935 (U/mL) and 373.941 (µg/mL), respectively.

Keywords: Classification; extraction; gel filtration chromatography; optimization; plant; protease

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