IGEM team meeting notes 4/28/14

iGEM planning 4/28/14 (copied from google docs https://docs.google.com/document/d/1B0lAqOm-4UO-KD4TWxLjOOPby98gLlW1dYFfDA2N1iM/edit?usp=sharing)

Topics for today:

• Bruce German at UC Davis - scientific advisor

• homework from last week, on which proteins to express

• fundraising
  • storyboarding fundraiser video
  • restart crowdtilt?
  • communication updates
    • Chr. Hansen will give us free samples.

• more casein experiments?
  • reforming micelles from powder (pseudo-milk)
    • change pH
    • change Ca
    • sonication

Seafile link: https://files.counterculturelabs.org/

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Glycosylation is the process of adding carbohydrates to protein.

Bruce German (UC Davis) seemed to think that glycosylation on the kappa casein play a role in the structure and/or formation of micelles. At davis they have an enzyme that can cleave off carbohydrates from the casein [is that deglycosylation]

With rennet you get a gel that forms, essentially a network of micelles. Small-curd cheeses-- e.g. cream cheese, cottage cheese-- do not require rennet, only acids, because they do not rely on this micelle network. We may want to attempt a cottage cheese with the bodybuilder casein, as a troubleshooting step if nothing else.

The name “casein kinase” is somewhat misleading, because casein is not the primary biological focus of these enzymes. Casein was the original test protein for these kinase enzymes, and the name has stuck.

We want to identify the specific casein variants we’re interested in; cross-referencing the Caroli paper (Table 2) with the MUSCLE multiple sequence alignment for bovine kappa casein would give us useful information.

What other papers are we still missing? There’s a few on artificial micelle formation I couldn’t get to

Very nice paper on “the real casein kinase”: http://www.sciencedirect.com/science/article/pii/S0968000412001910

Is this likely to be the only kinase acting on casein? What abou the Ca/calmodulin dependent vs independent kinase activity? Let’s read the paper off-line…

Patrik can you summarize why the genetic variants of caseins are of interest? To optimize health benefits?

Bovine kappa casein has 11 described genetic variants, A and B are most common. B is associated with better coagulation properties, and some potential health benefits, so that’s the one we should focus on in my opinion

Human kappa casein has 4 different forms; no idea on health effects

I sent out links to MUSCLE alignments for 17 bovine, and bovine + 4 human variants: http://www.ebi.ac.uk/Tools/services/web/toolresult.ebi?tool=muscle&jobId=muscle-E20140428-205013-0545-63725690-pg http://www.ebi.ac.uk/Tools/services/web/toolresult.ebi?tool=muscle&jobId=muscle-E20140428-233001-0975-29375072-oy The job results will be available for 7 days.

Still want to correlate SNPs in the casein genes with 23&me data, and any health effects described in human genomics literature (and GWAS data?)

Craig: if we take the longest gene and clone it, it will likely include all the splice variants

Patrik: we want to express the active version of the protein

Juul: I personally don’t really care about creating biobricks. I care about creating cheese. Who cares about creating biobricks?

Do we aim to package expressing alpha, beta & gamma caseins all into a single construct, or do each one individually.

Craig will look into pricing gene blocks from IDT for next week.

http://webecoist.momtastic.com/2011/01/11/from-another-udder-nine-nutty-non-cow-cheeses/ I know about this. It is currently illegal in the european union so you have to get it from the black market.

Storyboarding - here's the shotbox link: http://beta.shotbox.se/stories/3082

Fran’s 15sec video was fun - maybe expand to a quick 30sec version, and put up on CrowdTilt?

Jing’s Kickstarter video outline: https://docs.google.com/document/d/1AF6vuKMHD0N79nNXOBwLjNGvtJcxpqi5bg-RYE5m01s/edit

Fran (& anybody else) will try to flesh out the shotbox over the next 1-2 days into a 30-40 second file - by Friday

Possible next experiments to attempt:

Treat the bodybuilder casein with EDTA and/or sodium to reduce the amount of bound calcium, to improve casein solubility pH - see casein solubility curve at http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/enzyme-reagents/casein.html Sonication Use of a high-shear micro blender Attempt cream cheese or another variety that doesn’t require coagulation

• NEXT STEPS:*

• 30 second video by Friday - Fran leads
• Patrik: contact sponsors; contact Bruce German
• Craig: break point for # genes to order from IDT?
• Patrik & Craig: identify first genes to order (K-casein B, CMP? Human K-casein, Beta-casein? Fam20C kinase?)
• LAST WEEK’S HOMEWORK, YOU LAZY ASSES! ;-)