Integrated in Silico and Experimental Approach towards the Design of a Novel Recombinant Protein Containing an Anti-HER2 scFv

Abstract

Biological therapies, such as recombinant proteins, are nowadays amongst the most promising approaches towards precision medicine. One of the most innovative methodologies currently available aimed at improving the production yield of recombinant proteins with minimization of costs relies on the combination of in silico studies to predict and deepen the understanding of the modified proteins with an experimental approach. The work described herein aims at the design and production of a biomimetic vector containing the single-chain variable domain fragment (scFv) of an anti-HER2 antibody fragment as a targeting motif fused with HIV gp41. Molecular modeling and docking studies were performed to develop the recombinant protein sequence. Subsequently, the DNA plasmid was produced and HEK-293T cells were transfected to evaluate the designed vector. The obtained results demonstrated that the plasmid construction is robust and can be expressed in the selected cell line. The multidisciplinary integrated in silico and experimental strategy adopted for the construction of a recombinant protein which can be used in HER2+-targeted therapy paves the way towards the production of other therapeutic proteins in a more cost-effective way.

Identification and Functional Analysis of Cytokine-Like Protein CLEC-47 in Caenorhabditis elegans

“…Normally, it is necessary to have a purified protein to determine the precise biological function of a novel gene that encodes a secretory protein. By utilizing a purified protein, it is possible to decipher the function of the protein directly ( 44 ). In this study, we observed that the expression of clec-47 is highly correlated with bacterial infections, which suggests a critical role of CLEC-47 in the host immune response.…”

Modelling hCDKL5 heterologous expression in bacteria

“…This list of reactions was filtered by removing those (non-core) reactions showing more than 30% variation between their minimum and maximum fluxes during a FVA as described in Material and Methods. Overall, we found 15 gene-encoded reactions displaying an increased flux in this condition: the majority of them (11) were involved in histidine biosynthesis, three in phenylalanine, tyrosine and tryptophan biosynthesis and one in riboflavin metabolism. We found a similar scenario (i.e.…”

“…Also, these computational models may help identifying the most suitable approaches to get to target (protein) overproduction. Indeed, it has been recently acknowledged that the most innovative approach currently available to improve the yield of recombinant proteins while minimizing wet lab costs relies on the combination of in silico studies to reduce the experimental search space [11]. Among all the available in silico approaches, genome-scale metabolic models (GEMs) offers the possibility to predict a cellular phenotype from a genotype under certain environmental conditions and, importantly, to identify possible metabolic targets to improve the production of valuable compounds, while ensuring sufficiently high growth rates [12][13][14].…”

How promising are HIV-1-based virus-like particles for medical applications

“…Concerning targeted delivery, our group combined an in silico study with an experimental approach aimed at the development of a targeting motif that associates the singlechain variable domain fragment (scFv) of trastuzumab, a monoclonal antibody against the human epidermal growth factor receptor type 2 (HER2), with the HIV protein gp41 (Santos et al, 2021). Firstly, computational tools were used to perform docking studies to ascertain which residue in gp41 was the most adequate to fuse with the scFv.…”