Posted on behalf of the Rookie Rocky

In the world of marketing, brands are arguably as valuable as anything else. The idea is that a distinguished brand promises the quality of service or goods that can meet the customers’ expectations.

In the world of scientific research, I wonder if it’s pretty much the same. Recently, I have been struggling to get a paper published, and it seems to take more effort to satisfy the reviewers and editors than ever before, even though I wrote the paper in exactly same way as I have for many years. The message I get is: now you can breeze through the process no more. The reviewers question everything and the editors seem to be a lot more cautious about their concerns.

As a result, I’ve started wondering whether I got an easy ride under the established “brands” of my PhD and Post-Doc supervisors. Would my work have been published had I worked for myself or someone just like me – a newly independent academic? To qualify this question, I think almost all reviewers and editors, of course including those excellent ones at Nature journals, are fair. They do not judge the merit of a manuscript by whether a novice or a Nobel laureate generated it. Good papers are good no matter who wrote it, which is something I really like about science. The trick comes when a paper is not that good, nor that bad – that’s when the marketing effects may factor in. An established scientist would certainly have a strong publishing record that backs his/her credibility, which is something that rookies have to earn. It is just like we founded a start-up company, and the first thing we need to do is to familiarize potential customers with our new brand. Continuing the analogy, it is quite reasonable that people would choose to buy Coke or Pepsi rather than Rocky-Cola. That is not something we should complain about. In some ways, I feel this is rather nice as it will provide a window to aspire to higher standards: Hopefully one day I will have my own brand that compares with those of scientists I admire, and Rocky-Cola will be something that people really enjoy.

I was particularly struck by a recent paper from the Kozmin group on the mechanism of action of bistramide A. This very interesting natural product demonstrates promising anticancer activity, but is also highly toxic when delivered to mice. Interestingly, two other members of this family of compounds known as bistramides D and K, which both lack an enone moiety that is present in bistramide A, have been shown to be much less toxic while maintaining promising levels of anticancer activity in a mouse xenograft model.

The mechanism of action of this family of natural products has been hotly debated, but a few years ago the Kozmin group identified actin as the potentially therapeutically-relevant cellular target. This hypothesis was strongly supported by this group’s recent discovery and high resolution X-ray characterization of a bistramide A/actin complex. However, the mechanistic role, if any, of the conspicuous enone moiety of bistramide A could not be determined from this structure because this portion of the crystal was highly disordered.

In their recent PNAS paper, Kozmin and coworkers harnessed the remarkable efficiency and flexibility of their previously reported total synthesis of this complex natural product to prepare a series of elegantly designed analogs that collectively revealed the criticality of this enone moiety for potent cell-based growth inhibition. Moreover, consistent with the X-ray structure, these studies demonstrated unambiguously that both the spiroketal and amide subunits of bistramide A are required for high-affinity non-covalent interactions with actin that can lead to the severing of actin filaments. Follow-up studies with mass spectroscopy and a synthesized fluorescent analog collectively demonstrated that the enhanced cell-based activity attributed to the enone is due to covalent modification of the target protein, likely via conjugate addition of a cysteine residue. Collectively, these results support a dual mode of action of bistramide A involving the severing of filamentous actin as well as covalent modification of this protein target.

Interestingly, these results reveal a potential explanation for the increased in vivo toxicity of bistramide A relative to its enone-lacking counterparts. The severing of actin filaments (which does not rely on covalent modifications) may be sufficient to inhibit the proliferation of rapidly dividing tumor cells, whereas the dose-limiting toxicity may be caused by enone-mediated covalent modifications of this ubiquitous protein target. This compelling hypothesis remains to be tested, but this paper clearly demonstrates the critical importance of fundamental understanding of small molecule function to guide the search for more effective and less toxic therapeutics. It also represents a striking demonstration of the tremendous power of an efficient and flexible total synthesis of a complex natural product to enable the execution of illuminating experiments that are otherwise simply not possible.

Marty Burke is an assistant professor in the Department of Chemistry at the University of Illinois in Urbana-Champaign. His research focuses on the synthesis and study of small molecules with the capacity to perform higher-order, protein-like functions.

Posted on behalf of Retread

The mass of the earth is given by my physics book (Halliday 6th Ed.) as 6 × 10^27 grams. If we made just one molecule of each protein containing n amino acids linked together, when would we run out of material? Make a guess. I found the results surprising.

Assume the earth is made of nothing but hydrogen, oxygen, nitrogen, carbon and sulfur. Clearly not true, but we’re going for what mathematicians call an upper bound. If mathematicians can get away with things like “consider a spherical cow” I can get away with this. (The cognoscenti may wish to go for a least upper bound). Proteins are linear chains of 20 different amino acids ranging in mass from glycine at 79 Daltons to tryptophan at 204. When linked together by an amide (peptide) bond, 18 Daltons of mass is lost (water is split out). So figure the average amino acid at 100 Daltons (roughly).

So there are 20 × 20 = 400 distinct proteins of 2 amino acids, 8000 with 3, 160,000 with 4, 3,200,000 with just 5. Shorties like this are called peptides (or polypeptides) and just when you start calling them proteins seems to be a matter of taste.

We’re figuring the mass of the typical amino acid at 100 Daltons, but a Dalton doesn’t have much mass. It is 1/12 the mass of a single atom of carbon-12, Avogadro’s number (about 6 × 10^23) of which have a mass of 12 grams. So one Dalton has a mass of 10^-24 grams (roughly).

The number of distinct proteins containing n amino acids is 20^n. The mass of each protein (in Daltons) is (roughly) 100 x n — depending on the amino acids chosen. The mass of the collection of distinct proteins of length n in grams is (20^n) x (100 x n) x (10^-24). It’s clear that we’re over 1 gram for the collection at only 24 amino acids (as 20^24 is much larger than 10^-24. How far over? 2^24 × 100 × 24 = 40,265,318,400 = 4 × 10^10 grams.

As noted, the mass of the earth is 6 × 10^27 grams. So we’re not too far away at 24 amino acids. Certainly no farther away than another 17 amino acids as 20^17 is much greater than 10^17.

So, the mass of the earth (which isn’t all carbon, hydrogen, etc… ) isn’t enough to make just one molecule of each of the possible proteins 41 amino acids long. 41 amino acids is a very small protein (some would call it a polypeptide). Just about every protein of biological interest is much larger. The champ is a muscle protein called titin which has 27,000+ amino acids.

So what? It means that chemists will never be able to explore more than a tiny morsel of the space of possible proteins. Perhaps computationally we will (I doubt it), but that’s the subject of a future post.