Re: How Do the Proteases Found in Papaya, Pineapple and Kiwi Tenderize Meat?

Hi Scott,

I am by no means an expert on these proteases, but I did some poking around the web and will share with you what I found. I think a couple of hours of library or web research on your part should be able to fill in many of the gaps in my answer.

Here are some summaries of the enzymology of the peptidases you mentioned:
Papain, http://www.chem.qmul.ac.uk/iubmb/enzyme/EC3/4/22/2.html, and http://en.wikipedia.org/wiki/Papain.

Bromelain, http://www.chem.qmul.ac.uk/iubmb/enzyme/EC3/4/22/33.html, and http://en.wikipedia.org/wiki/Bromelain

Actinidin, http://www.chem.qmul.ac.uk/iubmb/enzyme/EC3/4/22/14.html

Ficin, http://www.chem.qmul.ac.uk/iubmb/enzyme/EC3/4/22/3.html

As you can see, these enzymes have similar but different EC (enzyme classification) numbers, which is an indication that they are very similar but have slight differences in their catalytic preferences. The subclass EC 3.4.22 is the cysteine endopeptidase class, so they are all classified as cysteine endopeptidases, which are endopeptidases containing a cysteine residue (sulfhydryl group) in the active site that is involved in catalysis. See http://en.wikipedia.org/wiki/Cysteine_protease.

It's not possible for me to give a short explanation of the subtle differences between the different endopeptidases (proteases). They all probably share the same overall structural fold, but will have amino acid sequences with conserved (shared) regions and other regions will be quite different. It should be relatively easy for you to look up their different amino acid sequences and possibly view some crystal structures as well, depending on your level of access to scientific journals. To get you started, here is a database entry for papain: http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?db=protein&val=387885. You can use the same NCBI website to search for other protein sequences.

One way to compare the similarities and differences of the different proteases, at least on the primary sequence level, is to do a sequence alignment. Collect the sequences, then enter them into an alignment program such as Expasy ( http://www.expasy.ch/tools/sim-prot.html). The different proteases are likely to have slightly different catalytic activities, stabilities, substrate preferences, etc.

It was not easy for me to find out the concentration of the different proteases in their "host" fruit, but I did find an old article that would allow you to infer an approximate value:
www.scielo.br/pdf/babt/v43n5/a08v43n5.pdf. I think you might find the general style of the article interesting to read, as you might find this one too: www.jbc.org/cgi/reprint/207/2/515.pdf.

The popularity of papain over the others is almost certainly due to availability and cost.

I wanted to comment a bit on your experimental idea. Ideally, you'd want to compare the tenderizing activity of proteases using the same amount of each protease. This will be very hard to do using fruit extracts, but it seems that it is possible to purchase commercial solutions of some of the proteases, at known concentrations.

You should take some time to carefully devise a "tenderizing metric" for your experiments. Will it be based on color change, on some mechanical change in the meat that you could quantify some way (yield to a weight?). Maybe you could shred some meat and mix in different proteases in different vials, in water. If you keep each solution stirred, maybe different levels of cloudiness could be quantified.

-Alex Tobias