Proteases and their inhibitors - a biochemist's overview of the what, where, when, & how

Today I thought I’d tell you more about types of proteases (protein cutters) and protease inhibitors. The kinds our bodies use, the kinds viruses use, and - of course - the kinds we use in the lab!

This post was inspired by the other days’ talk about Pfizer’s new covid pill, Paxlovid, which is an inhibitor of the coronavirus MPro protease - way more on that here: blog (also has some figures): https://bit.ly/paxlovidMPro YouTube:    • The basics & biochemistry of Paxlovid...  

That’s a specific protease inhibitor inhibiting a “picky” protease. But in the lab we often use generic protease inhibitors that inhibit promiscuous proteases to keep them from chewing up proteins we’re trying to purify. But we also often take advantage of proteases! Let me explain…

for long text, see blog form: http://bit.ly/proteaseinhibitors ⠀
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jargon note 1: “Enzymes” are usually proteins (sometimes protein/RNA complexes (like the case with ribosomes) or just RNA (we often call such enzymes ribozymes) and they mediate and speed up (catalyze) reactions by doing things holding the molecules together in the right positions for whatever they need to do and providing an optimal environment for the reaction to take place. ⠀
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jargon note 2 - “protease” and “peptidase” are often used interchangeably but technically a peptidase specializes in cutting short chains of amino acids, whereas a protease specializes in cutting long chains of amino acids (which in their folded form are better known as proteins). ⠀
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We can classify proteases/peptidases in a couple of ways…⠀
where they cut: “Endo”protease/peptidase refers to ones that cut in the middle of peptide chains and exoproteases/peptidases chew off the ends. ⠀
what cutting mechanism they use (what gives these scissors their blade): ⠀
pepsin is an example of an aspartic protease - it uses an Aspartate (Asp) amino acid residue to help water attack and break a peptide bond. Glutamic proteases work similarly, but they use Glutamate (Glu) instead of Asp. ⠀
metalloproteases use a metal to help the water out⠀
In all of those, water is doing the actual work, so there’s no “covalent intermediate” whereby part of the cut protein is stuck to the protease. However, with serine proteases (which I will discuss in the most detail) which use Serine (Ser), there is a covelent intermediate. Similarly for cysteine proteases (which use Cys instead of Ser) and threonine proteases which use Thr instead. So it’s protease + peptide → proteasepep + tide → protease + pep + tide (hopefully that’ll make sense later and/or in figs)⠀
another terminology note that’s easy to get confused about - when we classify proteases as “aspartic proteases” or serine proteases, we’re talking about what amino acid is in the active site of the protease doing the work. We are not referring to where on the protein or peptide the protease likes to cut. (i.e. a serine protease doesn’t prefer to cut next to serine). I wanted to put this here because I’ve heard people think this, which is totally understandable because when we talk about serine phosphatases we are talking about enzymes that remove phosphates from serine!⠀

for more on the mechanisms & uses see: http://bit.ly/proteaseinhibitors

And, I will probably make a video going deeper into the mechanisms