Crystals of proteins

From complexity to the extreme order

Greta Faccio, PhD
4 min readMay 24, 2020

Crystals are not as uncommon entities as we might think. They hold a great value for catalysis, imaging, and drug delivery applications. We might be used to see water crystals in our ice cream, but we can also find them in our makeup, and in our fresh or aged cheese.

Those white crunchy spots looking like small stones we find in Parmigiano reggiano or Grana Padano are not mold but crystals made of lactic acid or amino acids. The formation of crystalline structures made of protein was observed already in the XIX century and food scientists know them very well even nowadays. Once avoided as imperfections in food, they can now add a crunchy touch.

Cosmetic formulation can also contain crystals, more often inorganic ones (Photo by Juliana Malta on Unsplash).

In a medical context, a protein very familiar to many of us and it is the only used in medical settings in the form of a crystal is insulin. The crystal form proved fast to act and more stable than any other form. It is widely used to treat diabetes since the 1930s and provided at an affordable price thanks to its recombinant production using biotechnological means. Insulin crystallisation was achieved in 1926. For reference that was the year Queen Elizabeth was born and Winnie-the-Pooh was first published.

Fun fact: protein crystals have been successfully grown even in space in microgravity on the International Space Station ISS reaching sizes above 1 mm. On the other side, we find them on the shelves of the supermarket if we look well thanks to their purity and the craze for protein-rich food.

Scientists have however been long interested in crystals made of proteins, and especially when formed by only one type of protein that assembles into highly regular structures. This type of crystals has been essential to elucidate the three-dimensional structure of the protein in it.

Three-dimensional structure of a human insulin analogue (PDB ID:4CXL)

Procedures aimed at the growth of these crystals started to be developed for the determination of the structure of proteins using X-ray in the 1930s. First they used proteins that could be easily isolated from natural sources. Later on a wider range of proteins became available thanks to the new know-how on what DNA was, how to manipulate it, and how to use it to produce specific proteins.

A prerequisite if often that the solutions containing the protein you want to crystallise is highly pure and the protein highly soluble, this means does not settle immediately at the bottom of the container or aggregates in clumps. An excellent open-access review article is available on the topic. You can buy the necessary to crystallise your proteins easily online and a low resolution microscope can help you check if crystals are forming. Crystals can have very different shapes from think needles to regular transparent cubes.Crystal growth can be encouraged by a high concentration of the protein in solution that pushes the protein molecules to interact with each other. Otherwise the deposition in thin films, stirring, and the reshaping while forming using a laser has proven very promising.The formation of the crystals follows precise geometries that are dependent on the protein itself and the conditions used.

Protein crystals and the future.

The crystals formed by proteins might be small and invisible to the naked eye but offer a highly organised structure with good stability and rich in different chemical groups, provided by the amino acids forming the protein.

Protein crystals are much smaller than this one and range from a few hundred down to about 20 micron in each direction but they have also such a regular structure. Photo by Krystal Ng on Unsplash.

Protein crystals offer also highly structure with chemical groups that can be used to prepare materials with additional functionalities. Although very compact, protein crystals can also be seen as sponges that present regular cavities in their structure. These can fit other proteins, metals, or small molecules with a pharmaceutical action. Crystallisation already improves the stability of the protein, but further modification by chemical cross linking, by glutaraldehyde for example, can also offer a hand. The chemical groups offered by protein crystals in such and ordered and regular pattern can be used to trap enzymes producing a reaction of our choice. This has been applied to lipase, an enzyme degrading fats, and the functionalized protein crystal has been assembled directly inside a living organism, the bacterium Bacillus thuringiensis. Porous protein crystals such as the ones formed by the protein Cry have been produced filled with antimicrobial agents, a.k.a. antimicrobial peptide. This molecular Trojan horse proved very effectively picked up by infected macrophages… thus fighting the infection form the inside.

Although protein crystallisation is now only one of many approaches to resolve the structure of proteins, their formation is now even better understood and better achieved than years ago. Their bio-compatibility, recombination production, and possibility of functionalization offer many pros and few cons to their use in the future in different settings.

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