Post-publication review and PLOS’ experiment with the Synthetic Biology Collection

October28

An inducible luxI system (iptg) to produce the AHL above a threshold Pt. Kadam et al. (2016)

The iGEM 2015 synthetic biology contest was an important one for us. It marked our first attempt at putting together a project from IISER Pune. But beyond the novelty for us, many things were different this time around (#igem2015). First off, no preliminary or elimination rounds.

Secondly, we (yes, some self-backpatting here) organized an India Meetup in the run-up to the Jamboree. And third, and interestingly, the journal PLOS One (Public Library of Science) decided to use this as an opportunity to launch the PLOS iGEM collection, as a sort of meta-list, connected to iGEM. They decided to also go the radical way- with post-publication review. Time will tell how this latter experiment works out. And naturally our team’s efforts are there. With a lot of hard work put in by Snehal Kadam well after the contest and some griding-the-article together by mining long-forgotten (1 year ago!) protocol books, and some frantic emailing and interviewing, we managed to pull it off. You can read it here “Mycobacterium Revelio: Characterizing and Modeling Genetic Circuit Components towards a Bacterial Detection Tool”. The first 10 authors are BS-MS undergraduate students. Manasi and Neha are PhD students.

Micron-Scale Biological Devices

July25

The advent of micro-fluidics has been a boon to research in biology and medicine. Already many such devices exist in the commercial domain reducing what were a plethora of flasks, transfer processes and reactions at the macroscopic scale (even with a few micro-liters) to something that works in nano- and femto liter volumes.
We are using the relatively simpler process of soft-lithography by optimizing patterns using phot

The bacterial mother machine devlice (with high autofluorescence) and the assembly of the entire device (Manasi Gangan)

oresist that can be exposed to UV and post-baking used for building channels, flows and growth-chambers for cells.
We have reproduced the ‘mother-machine’ for bacterial cells described first by Wang et al. from Suckjoon Jun’s lab.

We are currently testing the results and developing some image analysis tools to extract meaningful numbers from the device.

For more on Biological Microfluidics, see the review by Velve-Casquillas et al. (& Phong Tran) 2010 Nano Today.

The new biologists fashion: Bacterial Physiology- Bringing back the old stuff

November27

The “post-antibiotic era” announced by the WHO, which was an update from April-2015 of an older report [1], suggests the need to understand bacteria is urgent as it ever was. We have been sailing the winds of Fleming from his 1928 discovery of Pennicillin-G from Penicillum notatum. For long chemists (led by the ‘magic bullet’ seekers) thought this to be some “biological”, “weak” and sill floundering. It took many other discoveries to demonstrate and industrialize production of antibiotics. But evolution is catching up [2]! The shocking part is not so much that it happened, but we were caught unprepared. The ups and downs in research notwithstanding, the need for research on the fundamentals is critical. One could argue for similar directed funding.

How do we know this is not some fear-mongering? Well, for one we now have a ‘bad-guy’. Strains of Klebsiella pneuomiae have been found to be resistant to all antibiotics known to humankind [3]. That is just the start. Mix it in with horizontal gene transfer and we have a real mess on our hands.

References:

[1] Anti-Microbial Resistance, World Health Organization April 2015

[2] WHO Warns Against Post-Antibiotic Era, Nature

[3] Superbugs from Wire.in

Bacterial cell division

November25
Populations of E. coli in microscopy

HupA (red) expressing E. coli with DIC (green) overlaid in confocal microscopy

The heat-stable protein HU was isolated first by Josette Rouviere-Yaniv and Gros in 1975 in a study where they systematically screened using an DNA-binding affinity-purification method for heat-stable proteins from E. coli cell extracts (1). This protein has been implicated in DNA replication initiation and cell cycle control (2,3). More recently the nucleoid binding HU protein from Mycobacterium tuberculosis
has been crystallized and the structure used to develop a small molecule which binds to HU and inhibits Mtb growth (4). This would appear to suggest surprisingly large number of physiologically critical bacterial proteins are still to be characterized that could help in the newly coined “post antibiotic era”.

REFERENCES

  1. J Rouvière-Yaniv and F Gros (1975) Characterization of a novel, low-molecular-weight DNA-binding protein from Escherichia coli. Proc Natl Acad Sci U S A. 1975 Sep; 72(9): 3428–3432.
  2. Jaffe A, Vinella D, D’Ari R. (1997) The Escherichia coli histone-like protein HU affects DNA initiation, chromosome partitioning via MukB, and cell division via MinCDE. J Bacteriol. 179(11):3494-9.
  3. Jason Kahn’s Lab (U. Maryland, USA) on Prokaryotic DNA Replication
  4. Bhowmick,T., Ghosh, S., Ramagopal,U.A.,Dey, D. Ramakumar,S. and Nagaraja,V. (2014) Targeting Mycobacterium tuberculosis nucleoid associated protein HU by structure based inhibitors. Nature Communications. 5:4124
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