Data reliability is a cornerstone of scientific work and the process to improve good practices for the evaluation and report of experimental results has been constant. Due to the extremely large variety of potentially alternative protocols, the identification of minimal sets of meaningful and implementable analyses suitable for recombinant protein quality assessment has been complicated. Further tests might become necessary when proteins are used for specific applications. As 1) protein quality evaluation should become a routine practice 2) specific applications require further analyses and 3) quality controls should be not perceived as a burden.

Membrane proteins are important drug targets (GPCRs, ion channels), yet are notoriously difficult to work with. We’ll present case studies on our latest research on the prominent drug targets CXCR4 and TRPV3 with direct extraction from crude cell membranes into lipid Salipro particles. This direct approach enables entirely new possibilities for de novo development and characterisation of biologics and small molecules, including phage display, B cell sorting, and cryoEM.

The STEAP1 is a cell-surface antigen over-expressed in prostate cancer, which contributes to tumor progression and aggressiveness. However, the molecular mechanisms underlying STEAP1 and its structural determinants remain elusive. Here, the fraction capacity of hydrophobic matrices on LNCaP lysates was evaluated followed by a Co-Immunoprecipitation (Co-IP) polishing. Several stabilizing additives were assessed, and the melting temperature (Tm) values ranked the best/worst compounds. A single step demonstrated higher selectivity of Butyl-Sepharose for cells impurities removal. Co-IP allowed to recover pure fraction of STEAP1. Tm of ~55ºC was determined and an a-helical structure was identified, confirming STEAP1 stability in the purification buffer.

This presentation discusses GSK’s refinement of the Lead Panel mAb generation stage as part of our revised End-to-End approach to keep the discovery funnel wider, for longer; to enable us to express and purify panels of antibodies in a high-throughput, semi-automated manner; to enable parallel prosecution of biological and developability screens, thereby accelerating our drug discovery process.

Due to the costly and time-consuming production of biological drugs, selective affinity purification methods are desirable. To achieve effective elution from the matrix when purifying by affinity-based methods, low pH is commonly used. These harsh elution conditions might cause the formation of aggregates and thereby greatly compromise the yield of the target protein, which can be a major problem for many biologic drugs. Therefore, combinatorial libraries for selection of ion-dependent high-affinity binders have been designed. These libraries and how to utilize them for selection and development of specific and efficient protein purification methods will be presented. 

Small synthetic compounds, tailor-made for selected biological targets, can be used as ligands in bioseparation. In this talk, we will show how rationally designed chemical combinatorial libraries support the development of robust peptidomimetics that can be easily adapted to several targets and to chromatographic and non-chromatographic methods.

In biotechnology, the use of a histidine tag that is molecularly fused to a target protein and forms a selective coordinative bond with traditional IMAC, NTA, or IDA functionalized materials to analyze and purify proteins quickly and specifically is well established. This talk introduces a new peptide tag that enables a novel affinity purification technique using underivatized silica.

The utilization of proteins to engineer materials has allowed the development of novel and advanced functional materials with great promise for broad applications. To guarantee the sustainable manufacturing of advanced protein-engineered materials, protein tag-mediated strategies that combine the purification and immobilization of recombinant proteins/peptides onto/into natural, synthetic, or hybrid materials in a single step are arising and attracting increasing interest. This talk will address progresses and challenges in this field.