eIF4E is part of a key group of proteins called initiation factors.

At Nanome, we’re dedicated to education. That’s why today we are starting a new series where we highlight the exciting fields of molecular exploration. In today’s piece, we take a closer look at protein engineering. But first, let’s take a quick look at proteins in general.

What Are Proteins?

Proteins cause most of the reactions that take place in humans, animals, and plants. They’re responsible for digestion, thinking, moving, fighting diseases, and more. Proteins are key for muscle development, as well as the regulation of body tissues and organs. Protein engineering is a multidisciplinary field, fusing computer science, biology, chemistry, and physics.

Proteins bring oxygen through the bloodstream, bring out energy from food, fire neurons and defend against invaders. Protein engineers today are on the cutting edge of biomedical research and development.

Amino acids are the building blocks of proteins, and amino acid chains fold into the three-dimensional structure of a protein molecule.

Now that we’ve explained proteins, let’s explore the world of protein engineering.

What Is Protein Engineering?

A protein’s structure determines its function. Protein engineering refers to the development and engineering of proteins that have new properties or functions. The genetic sequences of different proteins may be spliced together in one example of protein engineering. These so-called “chimeric” proteins end up having the properties of both.

As a general rule, modifications to the interior of the protein structure will likely produce deleterious effects on protein function compared to similar changes to surface locations. This is because amino acid residues in the protein interior are tightly packed. Changes here could result in steric hindrance and thus a change in the overall 3D structure.

Even the slightest modification of an amino acid sequence is considered protein engineering. For instance, a gene modification so that one or more codons code for a different amino acid type, insertions and deletions of codons or chemical modifications of the protein.

Engineered proteins fight disease. For example, scientists created Zinc Finger Nuclease (ZFN) to combat HIV. ZFN is an artificial restriction enzyme; that is, an enzyme that splits DNA into fragments at or close to certain recognition sites inside of a molecule.

Proteins that have been engineered are also treating cancers, autoimmunity/inflammation, infections, and genetic disorders, as well as adapting enzymes to the desired requirements of scientists.

Protein engineering will likely help test hypotheses throughout all aspects of molecular and cell biology, as well as aid the chemical, biotechnology and pharmaceutical industries by improving methods of protein-based product design.

Now that we’ve talked a bit about protein engineering, let’s look at the strategies that scientists deploy to engineer new proteins to combat disease and fill other demands.

What is Protein Folding?

The study of protein folding, for example, is a type of protein engineering, focuses on how proteins arrive at their native state (its properly folded/assembled, operative and functional form). A protein’s native state is determined by its amino acid sequence.

Now that we’ve looked at some of the protein engineering strategies deployed by scientists, let’s look at some known examples of protein engineering at work.

Protein Engineering Strategies

Proteins are engineered in two ways: via site-directed mutagenesis or random mutagenesis. Protein engineers often deploy rational protein design, directed evolution or both simultaneously.

A scientist employing rational protein design uses data and knowledge of the structure and function of a protein to make coveted changes to a protein by guiding its amino acids. Rational design enables protein engineers to do the following:

Create artificial proteins, which can then be applied to treat illness.

Research enzyme activity in living beings

Engineer genes

Create defenses against biological weapons

In directed evolution, random mutagenesis is applied in a controlled environment to a protein while a selection regime chooses those permutations with coveted traits. Directed evolution is the more popular strategy because rational design is only possible with structural knowledge of the protein.

In site-directed mutagenesis, one must know the structural information of the protein of interest. Random mutagenesis is best when solving complex problems or when structural information of the protein is lacking.

Novo Nordisk, for example, changed properties of enzymes (substrate specificity, thermostability, and stability) through site-directed mutagenesis.

GFP in Nanome

What Can You Do With Protein Engineering?

Designer Enzymes

Chemists from UCLA and the University of Washington engineered “designer enzymes” in 2008. The experiments were led by Kendall Houk’s UCLA chemistry group and David Baker’s Washington group with support from The Defense Advanced Research Projects Agency (DARPA). The designer enzymes they engineered could defend against biological warfare.

“The design of new enzymes for reactions not normally catalyzed in nature is finally feasible,” Houk said at the time. “The goal of our research is to use computational methods to design the arrangement of groups inside a protein to cause any desired reaction to occur.”

Enzymes are strong catalysts, according to co-author Jason DeChancie, an advanced chemistry graduate from UCLA who was working with Houk’s group. “[W]e want to harness that catalytic ability. We want to design enzymes for reactions that naturally occurring enzymes don’t do,” he said. “There are limits on the reactions that natural enzymes carry out, compared with what we can dream up that enzymes can potentially do.”

Now that we know a little bit about designer enzymes, let’s take a look at a few tools used by scientists to engineer proteins.

Check out Nanome highlighting molecules and proteins in Molecule of the Month

Protein Engineering Tools

The Low-Temperature Liquid Chromatography System

John Engen, a professor of chemistry and chemical biology at Northeastern University, and the liquid chromatography expert Waters Corp designed a low-temperature ultraperformance liquid chromatography system paired with a mass spectrometer. The purpose is to characterize proteins for the purpose of drug development.

The system resulted in an instrument called the nanoACQUITY UPLC System with HDX Technology. It features a refrigeration box that enables chromatography at 0 °C so that researchers can understand proteins through a process called hydrogen-deuterium exchange — or HDX — mass spectrometry.

Waters released the low-temperature HDX system in 2011, and now Thermo Fisher Scientific and Bruker, among others, offer HDX systems. “It took a few months to perform an HDX experiment” one decade ago, Engen says. “Now it takes a few days.”

An interesting note: Engen leads a Waters Center of Innovation meant to advance protein separation technology. Ohio State University chemist Vicki Wysocki’s Center focuses on mass spectrometry to distinguish protein complexes for the development of drugs.

Triple Quadrupole LC/MS system

Agilent Technologies launched in March 2017 a research-grade triple quadrupole LC/MS system. The idea is to expand research capabilities and sensitivity for environmental and clinical applications, as well as peptide quantitation, and forensic toxicology.

InvitrogenTrueCut Cas9 Protein v2

In November 2017, Thermo Fisher Scientific launched InvitrogenTrueCut Cas9 Protein v2. This next generation CRISPR-Cas9 protein was designed for maximum editing efficiency in broad range cells, like standard, immune, primary, and stem cells.

The Future of Protein Engineering

Market Overview

The growth in the protein engineering market is driven by several key factors. Rising government funding for protein engineering will be the main contributor to the market growing to USD 3.09 billion by 2025.

One such government initiative, the U.S. FDA’s critical path initiative, promotes the inclusion of advanced technologies into the drug discovery processes. Thanks to government initiatives, research activities, programs, and funds for research and development are on the rise in the protein engineering space.

As these above-mentioned cures, experiments, and tools demonstrate, the adoption of protein drugs over non-protein drugs is leading to advances in drug development. So, what does it all mean?

In a 2018 article titled “Protein Engineering May Be The Future of Science”, Bloomberg detailed how scientists are investing a lot of time, effort and money studying proteins.

According to the report, scientists are learning to design new proteins towards certain ends, whether that be new proteins for drugs and vaccines or cleaner ways of creating chemicals and other new materials.

David Baker, director for the Institute for Protein Design at the University of Washington, said that thirty years ago senior scientists discouraged his pursuit of protein engineering. Today, the field is alive and well. (Baker created a game called Foldit, wherein everyday people could try and determine how certain proteins would fold)

Degrado’s UCSF lab is studying how to create more stable medicines, as well as Alzheimer’s disease and similar neurological conditions affected by brain proteins. Baker’s lab is developing a vaccine to protect against all strains of the flu.

“[Protein engineers] can transcend the natural protein universe,” William DeGrado, a chemist at the University of California, San Francisco, told Bloomberg.

New Technologies And Softwares

Such funding is leading to new technology and software that is pushing the boundaries of protein engineering. One new, exciting technology affecting such change is VR. Viewing proteins at the nanoscale in three dimensions — and in such a manner so as they can be manipulated — provides scientists with new perspectives of the nanoscale. We covered how computers (including AI and machine learning) are aiding drug discovery more than ever.

Here is Nanome, our virtual reality-based molecular visualization tool is being used by top labs on multiple continents. If you’d like to learn more about what we do, visit us today.

Or, if you want to see scientists checking out molecules (including proteins) for the first time in virtual reality, check out the video below: