Protein Squinting

What’s the use?

Don’t dismay!  The structure of amino acids will help you infer!

                Determined by the sequence from different species, like primates, like bacteria, 2 proteins can perform the same function, an evolutionary relationship? An evolutionary relationship! The implication of this individual functional unit we find in nature, complex sequencing, beyond its normal function, pro-proteins, an extra sequence, like glycosylation, or a cell that says ‘no’ outside the cytoplasm, like an anti-body, recognition? Edman degradation fragments indirectly, routinely sequence, rapidly, DNA, decode into the protein sequence.  Reiteration, if the functional sequence spits out initially, can we tell post translationally?  Some side-chains cannot be changed at the end of the day, you can still check by mass spectrometry if this is ‘good enough.’ The Gene, The Proteome, the functional aspect of the nucleic acid, the origin.

                Nature acts to protect us.

              

                Stimulating anti-body production (get-away-from-me)

                The epitope is the part of the protein sequence that is going to represent the antibody, politically.

             

   Heavy-chain, light-chain, everything is covalent, produced from 4 separate polypeptide chains, the tertiary structure, a why-like shape, it doesn’t function in recognition at all, it helps the binding interaction take place.  Zoom in on that structure so you can see where it binds.  Red and Blue chains, both peptides are present in this domain, my FAV.  Flexible domains have more than two events take place. How do we make use? Immunity? We can form risky antibodies in your own rabbit, Polly Clonal. The rabbit (Polly) gets injected as you were once boosted.

We don’t know what’s appropriate.

Nature can do this efficiently.

The antibodies are built up in Polly’s blood. Higher-order species have developed many efficiencies.  Centrifuge serum, anti-serum, anti-bodies, all anti-gens to which Polly’s been exposed, they just exist there, figure that out.  Pictured up here are Polly Clonal’s rabbit anti-bodies, differently colored epitopes, red triangles, blue squares, green circles, a non-uniform (racemic) mixture, heterogeneity, why would we use these?

Pictured down here are Mona Clonal’s antibodies.  We’ve selected one predictable dose and response with different affinity. Most cells that produce antibodies have a very short lifetime.

Cancerous yeasts are immortal.  They don’t die.

Multiple myeloma is what we take advantage of.

We generate Polly Clonal’s antibodies, red cells, these will die, these (others) live, hybrids…Just blue, just red, blue and red together, the hybridoma cell line, one type of DNA, one type of antibody.  Polly Clonal is different from Mona Clonal.

Why do we care?

We can make use of precise analytical or preparatory chemistry, very small amounts, quantification, a read-out, fluorescence spectroscopy, affinity tag, the way that would work: a column format chromatography. Detection fits in with tracing.

E.L.I.S.A.: Enzyme-Linked ImmunoSorbent Assay.

The connection between enzyme and an antibody.

Signal detection? Tracer enzyme. Link. Better. Smaller. Direct. Backwards. If it sticks, we have a catalyst. Schematically what’s happening? Red dots are antigen, stuck to the walls of the vessel. Wash. Introduce antibodies. Wash. Introduce enzyme. Wash. Use the laser-dick. (A lot of steps are required to get it up.  That’s why we have designed Elisa. She’s good to us. She’s easy. Routine. Less work.  Sold easily. My precious…) Sandwich Elisa, she recognizes 2 antibodies, same antigen.  Make an elaborate assay, spectral readout, western blotting liked to electrophoresis (SDS-PAGE). You think that one of these bands is a specific protein? Radioactivity tag/auto-radiogram or Elisa, or fluorescence can also be used. That’s it for antibodies.

Cubans are pretty good synthetic chemists.  Not as good as nature. Nature puts out (more than Cubans?). But you can play tricks on nature that nature doesn’t do, unnatural substitutions. Synthetic peptides, short stretches of proteins can be made in a day, like a plant function, formylly, with Formylmethionine.  Higher-order mammalian (live-birthing teet sucker) cells intentionally make polypeptides. Vasopressin. ADH. Increase or decrease urination? Whatever’s functionally useful! Induce for treatment, roses full of loaded thorns. Desamino (take away) remove N term, split. We can do tricks like this to what was otherwise a natural material.

“Quaedam remedia graviora sunt quam ipsa pericula.” – Seneca

(Certain remedies are more grievous than the dangers themselves)