Making patterns with click chemistry

1.      Group of Fox used Tetrazine-trans-cyclooctene reaction for creating diffusion controlled 3D patterns (put it simply: particles) that could be filled with cells. Not a surprise cells stayed viable. The particles after all were made of hyaluronic acid derivatized with bioorthogonal chemistry that potentially can’t cause a harm. This work can find it’s future application in cell and tissue printing.

2.      Conceptually similar to the previous work, researchers in the group of Bart Jan Ravoo printed set of chemical functionalities using inverse-electron demand Diels-Alder chemistry (iEDDA) on the glass surfaces. In order to create those they employed a microcontact chemistry which enables to create large area patterns with high resolution. Essentially, you apply an ink that contain chemicals that you want to print on the surface on the PDMS stamps of defined geometry. 

Next, you apply the stamp on the functionalised glass. The chemical coupling will occur only at the sites where groups are close to each other. Very cool and straightforward. I believe that this sort of chemistry will find its further development in the field of biosensors and microfluidics as long as they fix a few limitations of this technique. This are the low reproducibility of the PDMS stamp-patterning (due to its elasticity) and a necessity to use apolar inks which is again due to the high hydrophobicity of the PDMS.

3.      A biomolecule patterning is also a passion of Dr.  Hongyan Sun whi published her work in Chemical Communication son the immobilization of peptides and proteins on surfaces with use of the iEDDA. A protein microarray technology may benefit from from this work.

Synthetic biology market is booming. I've just noticed that in the city next door (Singapore) anew SB-organization with wide profile (education, business, organization of conferences etc) just has opened. For those who are interested, check it out: http://synbiobeta.com/

Metallosensing DNA structures

Exciting and totally unexpected knowledge keeps me reading the scientific papers (apart from professional interest). This time I came across an article in Angewandte Chemie with a crystall structure ofa  DNA fragment (so far nothing new) with... T:T pair (non Watson-Crick) that is mediated by Hg ioins!

 

It looks like the metal perfectly held the B-structure of a DNA. In principle, this concept could be used in Metal-sensing nano-devices.

Dual color labelling in cell via SPIEDAC

Just read a fresh paper from Edward Lemke group in EMBL in last issue of Angewandte Chemie. They have 'adjusted' the SPIEDAC (Strain promoted Inverse Electron Demand Diels–Alder cycloaddition) such that now thay can efficiently dual label proteins in cell that bear byclononyne (BCN) and transcyclocotyne with a mixture of tetrazine and methyl-derivative of tetrazine containing dyes.

 It was well known that methyl-derivatized tetrazines are more stable and less reactive. I did not expect though that it will be so much less reactive so it will not be coupling with transcyclooctyne (see the picture). Great job!

Breakthrough in industrial mining of Uranium from... a see water!

Do you know that see water contains heaps of Uranium (in form of UO2++ ion) with average concentration 13.7nM. Well, it looks not impressive but water in the ocean in total have 100o times more of uranium that our earth crust. Problem is that no one came up with an idea how efficiently extract this precious metal out of water. Also, remember that presence of a similar type metals such as Vanadium, Copper etc which are presented in a water at even higher concentration will be outcompeting Uranium. And just to make task even more daunting, carbonate complexate with uranyl ion what leaves only 10(-17)  Mollar of free ion. Thus someone need to design a molecule with at least femtomolar affinity 10(-15) in order to effectively extract Uranyl at this negligibly low free concentration.
Group of Professors Luhua Lai (Peking University) and Chuan He (Chikago University) engineered a protein with fM affinity for Uranyl ion! They just have published in recent Nature Chemistry issue. The collaborative workflow they have presented in the paper is tremendous!  Briefly, they computed around 13000 of PDB known scaffolds for mutants that possibly can bind Uranyl which 'ccordination' geometry is very well known. They came up with 5000 hits, after couple of rounds of refining the library they were able to pick around ten with only four proteins amenable for expression. The best candidate has an affinity 37nM wich is far from perfect, but good luck and some of computation tricks led them to find a mutant with 1000-higher affinity (fM!). They were able to extract up to 95% of Uranyl from synthetic see water leaving just 3 ppb (!) of Uranyl in the filtrate. Now they are thinking of cheap device that would be able to replenish itself with protein Uranyl-binder. The most simple would be to use bacterias who can express this protein on the surface. So just by simply growing these bacterias now one can extract uranium out of see water. Although a whole idea is very smart and environmentally friendly (simple filtration of ocean water), use of bacterias will require stringent stages of water purification after it's been in contact with biomass given the huge masses of water that will be passing though these 'filters'.

News from conferences

Following the last post. Here is great list of recent reports in protein science and engineering field.

Raising genes from the Last Common Ancestor demonstrates its complexity

I do not why but papers about evolution always fascinate me (if they are not overwhelmed with population genetics and other hard core stats and math). This time I cam across a paper in JACS where authors used a computational technique called Ancestral sequence reconstruction (ASR) to rebuilt  ancient enzymes that nowadays form closely related bi-enzymatic complex. Basically, knowing the phylogeny (map of how species relates to each other) let you estimate the probability (maximum likelihood) that some mutation would occur back in its evolutionary history or vice versa: you can estimate the probability that current phylogeny would evolve given that ancestral protein sequence. Exactingly, you can go as back as you can (means - to the very Last Common organism Ancestor that we all evolved from, LUCA). In order to get that far the wider phylogeny tree you have the better: say from the most distant archaea to the bacterias. Authors asked a question whether LUCA back than 3.5 billion years ago had elaborate enzymatic networks. For that reason they rebuilt bieznyme complex (cyclase subunit HisF and the glutaminase subunit HisH) of imidazole glycerol phosphate synthase. Note that, without one another subunit would not work due to their close relationship in the synthesis reaction. They were able to show that the reconstructed proteins still retain almost the same specific activity and surprisingly able to tightly associate with each other. This work demonstrates that even LUCA had enzymes that were very closely associated and were able to perform such things as substrate tunneling (when product of one reaction is directly passed to another enzyme) or allosteric regulation (when product of one reaction regulates another enzyme). I wonder how would ribosome would look like given its high evolution conservativety.

The reason I decided to make a post about this paper is that ASR technique nowadays on the realm of very cheap gene synthesis let us 'play' with protein sequences such that we can go back in evolution and make probably more promiscuous enzyme that we can more easily 'teach' to perform reaction we want as these. Alternatively, we can improve folding properties of our protein by more targeted mutagenesis (since we have a good guess about its evolutionary history). Also, we might be able to produce an orthogonal protein/protein networks that still retains the specific activity while being not regulated by intracellular proteins. Any other ideas?