Recombinant expressions of sweet plant protein mabinlin II in Escherichia coli and food-grade Lactococcus lactis

Abstract

Sweet plant proteins, which are safe, natural, low-calorie sweeteners, may be suitable replacements for sugars in the food and beverage industries. Mabinlin II, a sweet plant protein, shows the most pronounced heat stability and acid resistance of any of the six known types of plant sweet proteins. However, mabinlin II is difficult to extract from the Capparis masaikai plant, which is itself becoming increasingly scarce. This limits the use of naturally acquired mabinlin II. In this study, recombinant mabinlin II proteins were expressed and purified in Escherichia coli and in food-grade Lactococcus lactis. Recombinant mabinlin II proteins MBL-BH (containing the B-chains of mabinlin II downstream fused with His-tag) and MBL-ABH (containing the A- and B-chains of mabinlin II downstream fused with His-tag) were expressed in E. coli in the form of inclusion bodies. They were then purified and renatured. The refolded MBL-BH was found to be 100 times sweeter than sucrose by weight, but it was not heat-stable. Refolded MBL-ABH was neither sweet nor heat-stable. Recombinant mabinlin II proteins were secreted and expressed intracellularly in food-grade L. lactis, in which the concentrated cell samples and culture medium samples were detected using enzyme-linked immunosorbent assay and Western blotting analysis with anti-mabinlin II polyclonal antibody. This study demonstrated that the single B chain of mabinlin II has a sweet taste. The recombinant mabinlin II proteins have been successfully expressed in food-grade L. lactis, which is a crucial step in the production of mabinlin II through microorganism expression systems.

Preparation and evaluation of anti‐renal fibrosis activity of novel truncated TGF‐β receptor type II

“…We attempted to obtain large‐scale quantities of soluble and active thTβRII from an E. coli expression system using the pET28a vector with thTβRII expression induced by optimal temperatures and IPTG concentrations. Unexpectedly, most of the overexpressed protein was insoluble, forming inactive protein aggregates referred to as inclusion bodies . We therefore developed a simple and efficient procedure for solubilizing and refolding the active form of thTβRII from such aggregates.…”

Expression of PprI from Deinococcus radiodurans Improves Lactic Acid Production and Stress Tolerance in Lactococcus lactis

“…The proteins in the gel were transferred onto a polyvinylidene fluoride (PVDF) membrane (Amersham Pharmacia Biotech, Buckinghamshire, England) and incubated with a rabbit anti-PprI polyclonal antibody (rabbit IgG, laboratory stock). Chemiluminescent signals on the PVDF membrane were visualized and quantified using a Gel Imager System (Bio-Rad Laboratories, Hercules, CA, USA) [ 12 ].…”

New Insight Into the Structure-Activity Relationship of Sweet-Tasting Proteins: Protein Sector and Its Role for Sweet Properties

“…Mabinlin is isolated from the seeds of Capparis masaikai Levl, which grows in subtropical regions within China. Based on its sequence, this sweet-tasting protein can be categorized into four members I, II, III, and IV (13). The mabinlin II (MW: 12,400 Da) consists of an A-chain of 33 amino acids and a B-chain of 72 amino acids, which are linked through two interchain disulfide bridges.…”