I’ve talked a lot about how scientists can try to figure out a protein’s structure (the way the protein’s atoms are arranged inside the larger 3D shape). But lost in the discussion has often been how proteins figure out their structure! So let’s step back and build from the amino acid to the peptide bond to the protein and how the biochemistry of the protein is the thing with all the sway at every level (primary through quaternary (I will explain so don’t worry if those terms sound scary)!

Today’s post text, after the vid, is adapted from some longer past posts. for more detail, see http://bit.ly/allaminoacidshttp://bit.ly/aminoacidstoproteins & http://bit.ly/peacepeptide & there’s a figure gallery at the end. Lot’s of ways to learn to love protein biochemistry!

Proteins are made up of long “polypeptide chains” of letters called amino acids linked backbone-wise through peptide bonds. There are 20 (common) genetically-specified amino acids, each with a generic backbone with to allow for linking up as well as unique side chains (aka “R groups” that stick off like charms from a charm bracelet). These chains fold up into functional proteins whose structure complements the tasks they carry out. They have several layers of structure that come from⠀

1. the the order of amino acids linked up in the chain (primary structure) ⠀

2. how the backbones of those amino acids interact through hydrogen bonds to give common motifs like α-helices and β-strands (secondary structure).

3. How the side chains interact to give you structure on top of that structure (tertiary structure) and then

4. how different chains sometimes interact to give you quaternary structure (not all proteins have multiple chains and only those that do have quaternary structure

You can get structure and not just “protein spaghetti” because the movement of peptide backbones is restricted by the nature of the peptide bond, which I go into in way more depth here: http://bit.ly/peacepeptide 

But basically, amino acids (like all matter) are made up of atoms (like individual carbons (C’s), hydrogens (H’s), oxygens (O’s), and nitrogens (N’s) are really small, but they’re made up of even smaller things – subatomic particles. These include protons, which are positively-charged, neutrons, which are non-charged, and electrons, which are negatively-charged. Atoms join together to form molecules by sharing pairs of electrons to form strong covalent bonds – 1 pair for a single bond, 2 for a double (which is stronger), and 3 for a triple.

Peptide bonds are a type of “amide bond” – they involve a carbonyl (-(C=O)-) attached to a nitrogen. And they’re special because the nitrogen, oxygen, and Cα (the carbon hookup up to the side chain) have to be aligned in the same plane in order for them to share electrons through resonance (electron delocalization), a phenomenon whereby atoms “donate” their extra electrons to a sort of shared pool. They “want” to participate in that because it helps stabilize them (you can think of it kinda like a play group where parents can help watch each other’s kids so there’s less burden on any one). So it’s “worth it” to stay stuck in a plane.

Therefore, when amino acids link up through peptide bonds, you end up with a sort of “chain of planes” where you can only have rotation at certain spots. And that rotation is further restricted by the nature of the side chain because of “steric hindrance,” which is basically a fancy way of saying, “dude – that’s my personal space!” Atoms (even though they’re super tiny) do take up space and space can only be taken up by one thing at a time – so the peptide can only twist a certain way if there’s room for its side chain to get situated without hitting anything. And bigger, bulkier things require more space.

Such structural limitations play a role in how proteins fold – chains of amino acids are called polypeptides, and these polypeptide chains can fold up into a few common “motifs” including alpha helices and beta strands that optimize backbone to backbone interactions (we call this “secondary structure”). Depending on their flexibility, they’re more or less likely to be found in different motifs. So, for example, Alanine (Ala, A) with the 2nd-smallest side chain (just a methyl (-CH₃) group) is commonly found in alpha helices, whereas Pro is not.

But steric hindrance isn’t the only thing determining where in a protein you’ll find the different amino acid residues. Even if it can be in a helix, is it out on the surface of a protein, near water? Or tucked tight in the protein’s central core? A major determinant of that is “polarity” and “hydrophobicity.”

Earlier we talked about how atoms – like those making up all of these different charms we’re discussing, link together through strong covalent bonds where they share pairs of electrons. The electrons in a covalent bond can be shared “fairly” or “unfairly” – fair sharing occurs when the partners have similar electronegativities (electron-hoginness), such as carbon and hydrogen, and it leads to an even charge distribution (non-polar). “Unfair” sharing happens when one of the sharers is more electron-hogging (electronegative) than the other – it’ll pull more of the shared electrons toward itself, leading to a partial charge imbalance we call polarity.

You often see such “polar covalent bonds” between oxygen or nitrogen (which are both highly electronegative) and carbon or hydrogen – the O or N steals more than its fair share, leading it to be partly negative and leaving its bonding partner partly positive. Opposite charges attract – even partial ones – so the partly positive parts of polar molecules like to hang out with the partly negative parts of other polar molecules (or other fully charged things).

Water is highly polar, so water molecules really like to hang out together. Thus, if you want water to hang out with something other than water you want that thing to be more attractive to a water molecule than another water molecule. If the water likes it (which happens if the thing is highly polar or charged), it’ll “dissolve” (get a full water coat) – we call such water-loving/water-loved things hydrophilic. Otherwise, the water will just “exclude” the thing from its network, leaving the excluded things to group together to make their surface area as small and hidden as possible. We call this the “hydrophobic effect” and it’s the main force behind protein folding – nonpolar amino acid residues are “hydrophobic” because they don’t have even partial charges to offer – so they fold up so that they’re in the protein’s interior, or at least facing away from the water.⠀more on the hydrophobic effect here: http://bit.ly/hydrophobiceffectPSA 

The extreme of this is seen with fully-charged amino acids. There are 2 side chains that are frequently negatively-charged at physiological (normal bodily) pH, Aspartic acid (Asp, D)(pKa ~3.65) & Glutamic acid (Glu, E)(pKa ~4.25). We call these acidic because they donate H⁺s & when they do they become negatively charged & now capable of accepting H⁺s (acting as a base) so we call them “conjugate bases.” It can seem kinda confusing because “acidic” residues often play important roles by acting as bases in their deprotonated form. The “acidic” refers to its neutral form being acidic.

There are 3 side chains that are frequently positively-charged & protonated at physiological pH – we call these “basic.” Lysine (Lys, K) (pKa ~ 10.28) & Arginine (Arg, R) (pKa ~13.2) are predominantly protonated at cellular pH, but Histidine (His, H) (5.97) is more “iffy” (that pKa tells you when HALF the groups are deprotonated on average so it’s not like you hit the pKa & bam they’re all deprotonated – you have a mix. Also, an important caveat is that pKas are context-dependent so the pKa you get from a table is likely close to but not exactly the “real” pKa in the situation you’re looking at). much more on protein charge here: http://bit.ly/ionexchangechromatography 

Protein structure can also be affected by post-translational modifications – things added onto the protein after the amino acid is put into the chain – such as phosphorylation (addition of phosphate groups which are bulky negatively-charged things) or glycosylation (addition of sugar chains). These alterations can make the protein have to rearrange to make everyone comfy again. 

Protein folding doesn’t always go right and sometimes it needs help. If a protein misfolds, machinery in the cell can try to refold it and if that fails, the proteasome can shred it up so aggregated (clumped-up) proteins don’t build up & become toxic to your cells (this is often a problem in some neurodegenerative diseases). But proteins can also get help during the translation process to make proper folding more likely. Basically, during translation, the amino acids are linked (w/help of ribosomes) 1 at a time to the end of a growing chain, going from N terminus to C-terminus. The protein chain starts folding as it emerges from the ribosome’s “chimney,” sometimes with the help of proteins called chaperones which can help hide hydrophobic parts that are coming out before the part of the protein that they’re ultimately going to “hide with” comes out. I talk more about this sort of thing here in my post on when you’re trying to get cells to make a protein for you (recombinant expression) and it’s not going well… : http://bit.ly/wherestheprotein

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