Chymotrypsin — the first protease structure

Today we have a guest post from Prof Stephen Curry, Professor of Structure Biology at Imperial College London.  Stephen blogs at Occam’s Typewriter and the Guardian and was one of the founding members of the Science is Vital UK campaign group.  He is currently visiting Australia and is giving a series of public lectures, the next in Sydney on 18th August.

Without further ado – here is Stephen’s post.

What does it look like?

This is a schematic representation of the protein chymotrypsin.

This image, made with PyMOL (using the coordinates from the Protein Data Bank entry 4CHA), shows the overall fold of the polypeptide chain of the molecule, along with a semi-transparent rendering of its surface. The lumpiness of the surface is due to the individual atoms of the protein because the structure has been determined at very high resolution.

What is it?

Chymotrypsin a protease molecule that participates in the digestion in the small intestine of proteins consumed as food. It acts like a molecular pair of scissors but instead of having sharp blades for cutting chymotrypsin uses chemistry. What this means is that it catalyses the cleavage of the peptide bonds between amino acids in selected targets. The three amino acid side-chains of the catalytic centre are shown as sticks in the picture above — from the top these are serine, histidine and aspartic acid. To the right of them you can see a deep pocket that  chymotrypsin uses to grab hold of a large, aromatic side-chain in a target proteins. During this transient embrace, the Ser-His-Asp ‘catalytic triad’ works, along with a water molecule, to destabilise and then break the adjacent peptide bond in the target. Repeated action of chymotrypsin and other proteases in the digestive tract breaks proteins down to their component amino acids so that ultimately they can be used by the body to build up new proteins, or as the starting material for synthesizing other essential molecules.

Where did the structure come from?

The structure of chymotrypsin — published initially by a team led by David Blow — was only the fourth protein structure to be solved by X-ray crystallography. It is a landmark because it was the first protease structure to be worked out by the technique and almost immediately revealed not just the architecture of the polypeptide chain but the detailed chemical mechanism that it deployed to break peptide bonds. The Ser-His-Asp catalytic triad has since been found in many other proteases in related and unrelated structures, providing intriguing molecular examples of convergent and divergent evolution.

Personally, the structure has a special meaning since David Blow was an inspirational figure who in helped me to get into protein crystallography at the outset of my career in 1989. I was fortunate to be able to return the favour many years later when my group solved the structure of the 3C protease from foot-and-mouth disease virus which turns out to be a distant relative of chymotrypsin.

β-Carotene, helps you see in the dark

What does it look like?

Image generated using Crystal Explorer 3.1 using data available from the Crystallography Open Database (# 2016501). Crystals are of a monoclinic P21/c structure with two molecules in the unit cell.

What is it?

Beta-carotene is a red-orange pigment found in many plants and fruit and is what gives carrots, pumpkins and sweet potatoes their bright orange colour. It was originally named by H. Wachenrӧder in 1831, who was the first to isolate and crystallise the molecule from the roots of carrots, though it may also be recognisable as E-number E160a, which it is assigned when used as a food colouring.

β-carotene is a natural precursor for vitamin A, needed by the retina of the eye in the form of retinal, which combines with the protein opsin to produce rhodopsin, a light absorbing molecule necessary for low light and colour vision. However, you don’t want to digest too much β-carotene, as overconsumption can cause Carotenosis, a benign condition which turns your skin orange!

Where did the structure come from?

Preliminary X-ray studies of β-carotene were conducted in 1937 and 1948, however, neither of these attempts were able to successfully resolve the systems molecular structure. It wasn’t until 1964 that that the actual atomic arrangement of beta-carotene was first determined (C. Sterling Acta. Cryst. 1964 17, 1224). The data for the structure presented here is from a much more recent study, from 2008, and is available from the Crystallography Open Database (# 2016501)

Keggin your pardon, ma’m – the structure of a heteropoly acid

What does it look like?

Dodecatungstophosphoric acid hexahydrate. Phosphorus atoms are shown in green, tungsten in purple, oxygen in red, and hydrogen in pink. Image generated using the VESTA (Visualisation for Electronic and STructual Analysis) software http://jp-minerals.org/vesta/en/

What is it?

A heteropoly acid is an acid whose anion (negatively charged part) is made up of multiple metal oxide units built around a ‘heteroatom’, such as phosphorus. This one is called dodecatungstophosphoric acid hexahydrate, with chemical formula (H₅O₂⁺)₃(PW₁₂O₄₀^3-). As you can see, this is a bit more complicated than your average acid, and the structure of the anion was not known until the 1930s, when  J.F. Keggin worked it out using X-Ray diffraction. This class of anions has since been known as Keggin ions.

The anions self-assemble in an acidic solution into the structure shown above. The phosphorous (P) heteroatom is tetrahedrally bonded to four oxygen atoms, which are enclosed by a tungsten oxide cage. The water molecules and acidic protons fill up the spaces between the anions. Various other elements can be substituted for P and W, giving ions with different structures and properties. They can be used as catalysts for processes such as the hydration of hydrocarbons to alcohols.

Where did the structure come from?

This udpate on Keggin’s original 1933 crystal structure was determined in 1977 by G.M Brown et al. and can be found in the Open Crystallography database (#1008010). They used a combination of X-ray and neutron diffraction, which is why the hydrogen atoms are so well resolved – neutrons are much better than X-rays at seeing hydrogen!