Monday, February 10, 2014

Protein A, G, A/G, L and Protein M


     Half jokingly I announced one day, "Scientists are normal people who are puzzled by things that most other people don't care about and chase its reality". Well its debatable, in terms of what really that should mean, I leave it for readers to justify. I scan several papers in a week, looking for important research papers, updates, through podcast and RSS feeds. But then usually the one that strikes me are the ones that relates to my routine practice. But every now and then i come up with something that simply blows me of the line, simply cause they are so predictable but haven't been thought about.

Fig 1: Protein A.
     Perhaps everyone is familiar with Protein A (Staphylococcal Protein A; SpA). It is a Type I membrane protein, a surface protein covalently bound to the peptidoglycan layer, which inhibits antibody mediated clearance by binding with IgG1, IgG2 and IgG4 Fc receptors. The structure of Protein A, has been studied in exquisite detail. It is postulated that Z domain (five homologous Ig-binding domains) which contains three alpha helices (arranged in an anti-parallel three-helix bundle), is the most important component that binds the immunoglobulin. The binding has been thought to occur either by 2 possible mechanisms. Rearrangement mechanism or Direct binding mechanism.

       S aureus uses the Protein A to chelate the antibody by binding to the Fc portion abolishing the activity of Fc, leading to Immuno evasion. This mechanism has been classically explained and forms a major part of Staph weaponry. Here's my punch point. A machinery that is too successful, is highly replicated in biology. That means, staphylococcus is not the only organism that produces a protein that can bind immunoglobulins. Protein A is usually used for research purpose to capture antibodies for purification purpose. A better version of this protein referred as recombinant variety, has better properties.

Other well described protein includes- Protein G, Protein L and the latest sensation- Protein M. I have summarized the important features of Protein A, Protein L and Protein G in table 1.

Table 1: Immunoglobulin binding Proteins.
       Protein G, expressed in group C and G Streptococcus with similar binding properties as that of Protein A. Protein G also binds to albumin, which is usually present in large quantities in preparations of blood. It has been modified to remove this additional activity, available as Rec Protein A/G. A domain in Protein G called as G B1 domain (or simply GB1 is known to render a protein soluble. This property has been made use of in research, by fusing insoluble Proteins with GB1 which is important for experiments such as NMR and crystallography. The recombinant variety has also better pH binding range (5 to 8.2).

          Protein L, differentiates itself from others by binding to Light chain (Kappa type) rather than the Fc portion. This property allows Protein L to bind to almost any type of immunoglobulin. It is isolated from Peptostreptococcus magnus, consisting of 719 amino acid residues.

It is important to note that the Proteins A, G, A/G and L are all available commercially for different types of purification based applications.

     So what has led me to write this post is the discovery of yet another molecule in the same pipeline, Protein M (from M genitalium). The surprise is that Protein M doesn't match with other binders in terms of sequence and structure. Instead of multiple Ig like binding folds, this has a large domain of 360 residues (with a very large binding area), binds principally to antibody VL domains. Unlike the Protein L which binds only kappa chains, this can bind both Kappa and Lambda chains. That has a lot of applications waiting to be used. But from bacteria's point of view, it has evolved a defense that is too good. The last I know, the protein is already in high demand to be commercialized like the others.

    Perhaps there are more antibody binding proteins out there in nature with different properties. We may need to look hard enough.
Mitsuru Tashiro, Roberto Tejero, Diane E Zimmerman, Bernardo Celda, Björn Nilsson, Gaetano T Montelione (1997). High-resolution solution NMR structure of the Z domain of staphylococcal protein A. Journal of Molecular Biology, 272 (4), 573-590 DOI:10.1006/jmbi.1997.1265

Sjöbring U, Björck L, & Kastern W (1991). Streptococcal protein G. Gene structure and protein binding properties. The Journal of biological chemistry, 266 (1), 399-405 PMID: 1985908

Akerström B, & Björck L (1989). Protein L: an immunoglobulin light chain-binding bacterial protein. Characterization of binding and physicochemical properties. The Journal of biological chemistry, 264 (33), 19740-6 PMID: 2479638

Grover RK, Zhu X, Nieusma T, Jones T, Boero I, MacLeod AS, Mark A, Niessen S, Kim HJ, Kong L, Assad-Garcia N, Kwon K, Chesi M, Smider VV, Salomon DR, Jelinek DF, Kyle RA, Pyles RB, Glass JI, Ward AB, Wilson IA, & Lerner RA (2014). A structurally distinct human mycoplasma protein that generically blocks antigen-antibody union. Science (New York, N.Y.), 343 (6171), 656-61 PMID: 24503852

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