Monkey cup protease for protein analysis

December 11, 2012 § Leave a comment

A monkey cup. Image from

A monkey cup. Image from

If “monkey cups” doesn’t get your attention, I don’t know what will. A recent paper in Molecular & Cellular Proteomics had this phrase in its title and, obviously, caught my attention. It turns out that the monkey cup plant, an insect-eating plant called nepenthes that has modified leaves that catch water from which monkeys drink, has a special protease. This protease may be useful for determining protein structures and mapping protein interactions, particular those of intrinsically disordered proteins.

David Schriemer‘s group at the University of Calgary came across this monkey cup protease when searching for an aspartic protease that was phylogenetically distant from standard workhorse protease pepsin. The Schriemer group is interested in hydrogen-deuterium exchange mass spectrometry as a method because it has the potential to reveal high-resolution structural and temporal details about complex protein systems. Proteins can be thought of as “breathing”; they exchange their hydrogens for deuteriums when bathed in D2O. The hydrogens that get switched for deuteriums help researchers figure out details of protein structure, folding and interactions.

Besides D2O, the method also uses pepsin to cut proteins into manageable pieces. But pepsin has its drawbacks. It isn’t always an efficient protease, and it doesn’t consistently cut all proteins at the right spots. “We want 100 percent coverage so we can track exactly where the deuteriums go,” says Schriemer.

So when the investigators came across the monkey cup protease called nepenthesin, they were intrigued. “We were surprised to find that nepenthes extracts are very poorly characterized, and only a handful of studies exist.  What evidence there was suggested it was worth a look,” says Schriemer.

The investigators grew a few nepenthes plants in their lab, fed them fruit flies to induce secretion of the plant’s digestive juices, collected those juices, and isolated the protease.

Schriemer and colleagues then tested nepenthesin in the place of pepsin. They found they could “now look at larger protein complexes and expect to get better sequence coverage,” says Schriemer. The protease “really extends the reach of the method and gets us closer to a proteomics-grade version of the technology.”

In particular, nepenthesin can better handle intrinsically disordered proteins than pepsin, which doesn’t deal well with the prolines and charged residues that often dominate these kinds of proteins.

Schriemer says the next step for the group is to scale up nepenthesin production. “We have many people asking for the enzyme!” says Schriemer.

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