Production and single-step purification of Brugia malayi abundant larval transcript (ALT-2) using hydrophobic interaction chromatography

Abstract

Abundant larval transcript (ALT), a novel filarial protein, has been shown to have great potential as a vaccine in the prevention of human lymphatic filariasis. In this study, we report a method for the production of recombinant ALT-2 protein, expressed in the cytoplasm of bacterium Escherichia coli in soluble form and purification in a single step using hydrophobic interaction chromatography (HIC). Fermentation was done by continuous fed-batch methodology with dissolved oxygen (DO)-controlled feed addition. The culture was induced with 1 mM isopropyl-β-D: -thiogalactopyranoside (IPTG). Up to 9 g/l dry cell weight (DCW) of biomass was obtained from 1.6 l of Luria-Bertani (LB) broth in a bench-scale reactor. Around 200 mg/l of purified ALT-2 with a yield of about 60% was obtained. This is almost a 2.5-fold increase in final protein yield compared to purification using immobilized metal affinity chromatography (IMAC).

Sophisticated Cloning, Fermentation, and Purification Technologies for an Enhanced Therapeutic Protein Production: A Review

“…HIC is used for both small and large scale purification, including hormones, and industrial enzymes (Roettger and Ladisch, 1989; Bhuvanesh et al, 2010). Several recombinant proteins such as anthrax protective antigen, human interferon alpha, etc.…”

“…Several recombinant proteins such as anthrax protective antigen, human interferon alpha, etc. have been expressed in E. coli were purified employing HIC (Gwinn et al, 2006; Salunkhe et al, 2009; Bhuvanesh et al, 2010; Wang et al, 2014). …”

Production of recombinant nonstructural 1 protein in Escherichia coli for early detection of Japanese encephalitis virus infection

“…The composition of the growth media is crucial for enhancing product formation as well as reduction of inhibitory compound formation (Manderson et al ., 2006; Tripathi et al ., 2009). Furthermore, one of the most popular methods to achieve high cell density is fed‐batch culture by controlling the nutrient feeding via pH, dissolved oxygen (DO) or specific growth rate (Lim et al ., 2000; Manderson et al ., 2006; Khalilzadeh et al ., 2008; Bhuvanesh et al ., 2010). Recombinant protein purification using the minimum possible steps is crucial to meet the required level of purity.…”

“…Recombinant protein purification using the minimum possible steps is crucial to meet the required level of purity. Affinity chromatography is versatile and can be used for improved purification of recombinant proteins (Bhuvanesh et al ., 2010; Tripathi et al ., 2010).…”

Hydrophobic Interaction Chromatography: Fundamentals and Applications in Biomedical Engineering

“…The purified peroxidase was extremely stable in different media, and therefore its commercialization seems promising. Bhuvanesh et al (2010) used a single-step method to purify a filarial protein (expressed heterologously in E. coli) with great potential as a vaccine for preventing human lymphatic filariasis. The purification method consisted of a HIC step.…”