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How can you engineer a protein for biotechnology research?

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Proteins are the building blocks of life and the key players in many biological processes. They can also be used as tools for biotechnology research, such as enzymes, antibodies, sensors, and therapeutics. But how can you engineer a protein for a specific biotechnology application? In this article, you will learn the basic steps and methods to design, produce, and test a custom-made protein for your research needs.

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Andrii Buvailo, Ph.D.

Science & Technology Communicator | Life Sciences
Palaniswami Rathanaswami

Biomedical Research Scientist, Consultant at PRSwami AbDev Inc. Instructor for Training Seminars on Label-Free…

1 Identify your target

The first step to engineer a protein is to define your research goal and identify the target protein that can achieve it. You may want to modify an existing protein or create a novel one from scratch. You may also want to enhance or alter some properties of the protein, such as stability, activity, specificity, or binding affinity. To do this, you need to have a clear understanding of the structure, function, and interactions of the protein and its potential partners or substrates.

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Palaniswami Rathanaswami

Biomedical Research Scientist, Consultant at PRSwami AbDev Inc. Instructor for Training Seminars on Label-Free Biosensor Tools in Biotherapeutic Discovery: SPR, BLI and KinExA.
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One of the important factor to identify for protein targets is where the target is present. It could be a soluble molecule or could be present on the cell membrane or inside the cell or in an organelle. Further it could be present in moving blood environment or in a tissue environment.

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2 Design your protein

The next step is to design your protein using computational tools and bioinformatics. You can use software to model the protein structure, predict the effects of mutations, and optimize the sequence for expression and purification. You can also use databases to search for similar or related proteins, domains, motifs, and sequences that can help you with your design. You can also use algorithms to generate new sequences or combinations of existing ones that meet your criteria.

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Andrii Buvailo, Ph.D.

Science & Technology Communicator | Life Sciences
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The area of AI-driven protein design is on fire! One example is Generate Biomedicines. Their Chroma platform learns Protein Data Bank and can be programmed to include specific features, such as symmetry, certain shapes, and functional elements, and it can understand descriptions of what a protein should do. Chroma includes a new type of neural network for 3D molecular data, a diffusion process that applies noise in a way consistent with protein structures, and an advanced technique for creating high-quality protein models. So, they can design novel proteins with desired shapes and properties. Another company Absci, uses zero-shot machine learning to design antibodies.

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3 Produce your protein

The third step is to produce your protein using a suitable expression system. You need to synthesize the DNA sequence that encodes your protein, either by chemical synthesis or by PCR amplification from a template. You then need to clone the DNA into a vector that can be introduced into a host organism, such as bacteria, yeast, insect cells, or mammalian cells. You also need to select the appropriate promoter, tag, and fusion partner for your protein to ensure its expression, solubility, and detection.

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4 Purify your protein

The fourth step is to purify your protein from the expression system using various chromatography techniques. You need to lyse the host cells and separate the protein from the cellular debris, contaminants, and other proteins. You can use different types of columns and resins that bind to specific properties of your protein, such as size, charge, affinity, or hydrophobicity. You also need to monitor the purity, yield, and activity of your protein using methods such as SDS-PAGE, Western blot, or spectrophotometry.

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5 Characterize your protein

The fifth step is to characterize your protein and evaluate its performance for your research application. You need to measure the physical and chemical properties of your protein, such as molecular weight, folding, stability, solubility, and aggregation. You also need to assess the biological activity and function of your protein, such as enzyme kinetics, binding affinity, specificity, and inhibition. You can use various techniques, such as mass spectrometry, circular dichroism, fluorescence, SPR, ELISA, or assays.

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Palaniswami Rathanaswami

Biomedical Research Scientist, Consultant at PRSwami AbDev Inc. Instructor for Training Seminars on Label-Free Biosensor Tools in Biotherapeutic Discovery: SPR, BLI and KinExA.
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Identifying the functional epitope of the protein is important to design an antagonist or agonist with best binding affinity and potency. Characterizing the epitope as linear or structural also help to express the protein in such a way to maintain the epitope to be used in invitro assays.

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6 Optimize your protein

The final step is to optimize your protein and improve its biotechnology potential. You may need to fine-tune your protein design, expression, purification, or characterization methods to achieve better results. You may also need to perform further experiments to validate your protein in different conditions, models, or systems. You can also use methods such as directed evolution, rational design, or combinatorial libraries to generate variants or mutants of your protein that have enhanced or novel properties.

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Palaniswami Rathanaswami

Biomedical Research Scientist, Consultant at PRSwami AbDev Inc. Instructor for Training Seminars on Label-Free Biosensor Tools in Biotherapeutic Discovery: SPR, BLI and KinExA.
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Optimizing the proteins to bind to its partner with specificity with out any non-specific binding to related molecules or homologous protein is important. Tighter the binding affinity would help to minimize COG.

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7 Here’s what else to consider

This is a space to share examples, stories, or insights that don’t fit into any of the previous sections. What else would you like to add?

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How can you engineer a protein for biotechnology research?

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1 Identify your target

2 Design your protein

3 Produce your protein

4 Purify your protein

5 Characterize your protein

6 Optimize your protein

7 Here’s what else to consider

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