Lecture 3 - trovenvolent bond.

secondary stsructure of protines. ribsomes make the unfolded proteins. proteins eventaully are going to fold. once they are folded they are going to retain the shape and not unfold unless u do something to the protein soltuions like hgih or cold temp or high acidicty. under physiolgoical conditions onces folded it stays folded. A proteins conformation is statbilized largely by weak interactions -

Stability of a proteins - is the tendency of a protein to maintian a native conformation - native conromatio - its biologically active confromation

chemica interactsion that stabilize these confromations - the strongerst is the disulfide bond which is a covalent bond between two sulfur atoms. this is the most uncommon one. the more common ones are noncovalent bonds. insulin has disulfide bonds. the non covalent ones are weak interactions. they are hydrophobic effect, hydrogen bonds, and ionic interactions.

hydrophobic effect is an effect. its not an intercition. hydrophobic molcules ten to cluster together . it is trough t obe the predominating factor for stabilizing protein confromations. they are all important but hydrophibic effect is the main driver of stability .

the entire protein is hydrophobic and if u force the protein into an aqueous enviorment. the water moelcules will form a solvation layer around the protien. the water mjoclules that are closest to the protein are highly ordered, the ones that are further away are not ordered. the ones that are further away are free to move around since they are not ordered. The solvation layer ones are hydrogen bonded to the neighboring water mocluels and they ahve low degree of feedome. this is thermodynamically unfavorable . the universe perfers high entropy system it will try to achieve entropy. but in this situtation there is order in the solvation layer, so u decrease entropy. this is unfavorable. for a real protein it will hid ethe hydrophobic amino acids inside and the philic ones outside to prevent forming ordered watermoclules. this is why the hydrophobic effect happens. one protein is folded it will retain the native conformation. most proteins are not rigid moclules, if u look at a real time u see they are not compelty rigid.

atp syntheis is highly dynamic and hemoglobin is alsodynamic but not as mcuh as atp . most proteins have soome level of movment.

how do prpteins fold into a 3d shape and what allows them to be dynamic (movment) instead of rigid(stuck). . .

talk about the peptide bonds its the carbon nitrogen covalent bond that links the two amino acids togther. the peptide bond actaully has resonance structure .it behaves like a double bond, double bonds are non rotabale. peptide bonds are not rotabale. if u were to measure the bond legnth of the carbon and nitrogen, it is slightly shorter than what u would expect a pure carbon nitrogen single bond be. this is also proof that this is a double bond .

segment of a protein -

the peptide bonds are non rotatable. so the carbon nitrogen bond does not rotate even when its folding. there are restrictions on rotations of the bonds. carbon- carbon bond between the carbyon and the lapha carbon are a true single bond. u can rotat the molceulce, the nitrogen to alpha cabon bond are freely rotatble. peptide backbond containes three types of covlaent bonds two are rotatble and one is not. proteins fold by rorating the two that can be rotated. the two bonds that can be rorated. we need to have a way. of descirbing how much roration is applied along the axis. phi angle is the dihedral angel taht concerns the bond between the nitrgorn atom and the alpha carbon atom. u can rorate along that axis and the degree of rotation can be -180 to 180. its the same as 360 menaing full roration. the psi angle the carbon and carboyl atom . the peptide bond is the W omega and 180 degrees for trans . the r and the O are opposite. that is the trans cofiguration.

not all360 degrees of ratiaotion is aviable. this is because there can be clashes within amino acids. as u rotate u can create clashes. so even tho u can rotate its not good. as u rotate as certain psi angels u can clash onto atoms. the unobtainle phi and psi angles dpepnds on what the next and previous amino acid is.

Ramachandran plot - it is going to visualize all the psi and phi angles in a protien. they take a value betweeen n-180 and +180. each black dot is a amino acid in a protien. it is the measuremnet of the phi and psi angels in a amino acid. 1 out of the 100 amino acids . not all foru nquadrants are qually occupries. u have a hig density of amino acids. vast areas that are empty because those phi and psi angles lead to clases. so the grouped ones are the most compatible.

seconday strcture is the lcocal psiatl arragnemetn. some will have only alpha or only beta sheets or only random coils. in alpha helices and beta sheets the phi and psi angles remain the same thoughout the segment. within an alpha helix, will have ismilar phi si angles. all amino acids

a helic will have a phi angel of -57 and psi of -47. alpha helix common protien secondary structure. linus pauling 1948 - we take alpha helixes and betta sheets for granted.

alpha helix backbone is wound around an imaginary axis. r groups protrude out from the backbone. each helical turn = 3.6 resides - 5.4 amstrung.

the pitch… 3.6 makes a complete turn and the elevation goes up by 5.4 A. the h bonds in an alepha ehlix is going to stabilize . if u look down the axis the r groups are protroucing outward. when u have a helical structure it needs to be left or right handed

intrahelical hydrogen bonds - every single electronegative atom that would be part of the peptide bond will be invovled in hhdrogen bonding. if its an alpj

So we already said there are a lot of hydrogen bonds

within an alpha helix. Okay. In fact, every single

electronegative nitrogen atom, so that would be

part of the peptide bond. and the electronegative

carbonyl-oxygen atom are involved in hydrogen bonding.

So each amino acid has two such groups.

So if it's an alpha helix that's made up of 20 amino

acids, that means there would be roughly 40 hydrogen

bonds within that alpha helix. Again, a lot of

hydrogen bonds. That's why this structure is stable.

If you look at the structure closely, the hydrogen

bond forms between the hydrogen atom attached

to the nitrogen atom of residue N and the carbonyl

oxygen atom of residue N plus 4, okay? Okay,

so amino acid number one would form a

hydrogen bond with amino acid number five.

Amino acid number two will form a hydrogen bond with

amino acid number six, and so on. Okay, then this

way, all amino acids are involved in hydrogen bonding.

Okay, so this is the main interaction that's

stabilizing the alpha helix structure. OK,

but there is sometimes an additional stabilizing

factor, and that's shown in this slide.

OK, this is again looking at an amino

acid or alpha helix along the axis,

straight down the axis.

OK, these numbers refer to amino acids. OK, now they're

simplifying this figure to, so each sphere actually

represents a single amino acid here. And you can see

in this view that amino acid number one in three

-dimensional space is closest to amino acid number four.

They're almost top and below of one another in such

a way that amino acid number one is closer to number

four than it is to amino acid number two. Okay?

And as you can imagine,

if amino acid number one has an R group that is

positively charged, whereas amino acid number four

has an R group that is negatively charged, because

these two amino acids are close in space, there's

going to be a favorable electrostatic interaction

between those two respective R groups, okay?

And this has a stabilizing effect

for the entire alpha helix, okay?

It's a double-edged sword, though. If amino acid

1 has a positively charged R group and amino acid

4 also has a positively charged R group, that's

going to destabilize that alpha helical structure.

okay and

let's say amino acid one is glycine amino acid number

four is also glycine what's special about glycine

it's r group is just a hydrogen atom okay it's

the smallest of the 20 amino acids okay they're so

small that they're not gonna interact with one another

right and in that case, the nature of the R group

does not stabilize or destabilize the structure.

Make sense?

All right, the next secondary structure element,

we have to start here, called beta sheets,

okay? But beta sheets are made up of beta strands,

so let's look at what beta strands are. It's just

a stretched out polypeptide chain, okay? But

it's not fully stretched out. As you can see here,

it has that zigzag shape, okay? So you can stretch

it out slightly more, okay? But it's pretty well

stretched out. So if you stretch out a polypeptide

segment like this, okay, you'll see that the R

groups protrude above and below that chain, okay?

Now, what are beta sheets? Okay, that's the

secondary structure element, second one we're talking

about. It's a collection of two or more beta

strands positioned side by side, okay? So, as I

alluded to earlier, there are two types of beta

sheets. At the top, this is an anti-parallel

beta sheet. Below, this is a parallel beta sheet.

The difference is the direction

of the individual beta strands.

Here, the top strand is going from left to right,

the next one right to left, and then left to right,

so it's going in opposite directions. That's why this

is an anti-barallel beta sheet. By the way, what

do I mean by direction of a polypeptide chain, okay?

It's just like, we talked about this.

Polypeptide chains have directionality.

You read from the N -terminus to the C-terminus

when you read the amino acid sequence, okay?

And that's the direction, okay? Left to right,

from N-terminus to the C -terminus. But if you look

at a parallel beta sheet, you can see all the

strands are going from left to right, okay? And

that's why it's called the parallel beta sheet.

Like alpha helices, the beta sheet structures are

also stabilized by a large number of hydrogen

bonds, which are shown by these dotted lines, okay?

Again, there are so many, okay, that

collectively they provide a strong force

that stabilize the beta sheet structure.

Okay.

If you compare a equivalent anti-parallel

beta sheet to a parallel beta sheet, okay,

so they are, they both contain three strands.

They're made up of the same number of amino acids,

also same number of hydrogen bonds altogether.

However,

in those cases, the anti-parallel beta

sheet is somewhat more stable than parallel

beta sheets, okay? So why is that?

The number of hydrogen bonds are the same, okay?

But one is more stable than the other, okay?

Remember for hydrogen bonds, there's

an optimal interaction length,

okay? That's very important between the

hydrogen bond donor group and the acceptor

group, but also the interaction or the

hydrogen bond angle is also very important.

Hydrogen bonds are strongest when the donor

atom, hydrogen atom, and the acceptor atom have a

180 degree relationship. They're linear. here.

So the way that the hydrogen bonds are formed in an

anti-parallel sheet, you have the maximally strong

hydrogen bond. In an anti -parallel sheet, the donor atom

and the acceptor atom and the hydrogen atom are

offset at an angle. This weakens those hydrogen bonds.

That's why anti -parallel sheets are

more stable than parallel beta sheets.

Third,

secondary structure elements you find

in proteins are called beta turns.

As the name implies,

this connects two ends of two adjacent segments

of an anti-parallel sheet. This is the

most common place that you'll find beta turns,

okay? So if you look at an anti-parallel beta sheet,

the first strand is going in this direction,

then suddenly it has to change in the opposite

direction, so it has to make a turn, right?

And that's where you often find beta turns.

It makes a 100-degree turn.

It's made up of just four residues, four

amino acids, okay? There are two types, type

1 beta turn, type 2 beta turn, you don't

need to be able to distinguish these, okay?

Whenever there are four amino acids that make

a 180 degree turn to the polypeptide chain,

you have a beta turn secondary structure there, okay?

And oftentimes,

well, the beta turn is stabilized by a single

hydrogen bond, shown here, okay? And often

you find glycine and proline residues as part

of the beta turns, okay? The beta turns are very

tight, okay? And it's favorable to have

glycine and proline to avoid any steric clashes.

Let's go back to the Ramachandran plot now, okay?

Because previously we discussed why there

are unoccupied spaces, vacant spaces, okay?

And we said that's because of clashes, potential

clashes. You have to avoid those clashes

so those phi-sci angles are not occupied.

But then why are they so highly concentrated

in certain regions? And now you see why. Okay.

Amino acids that are part of both

anti-parallel and parallel beta sheets

tend to have phi and psi angles that

are concentrated in this region.

Okay.

Right-handed alpha helix, they will have phi and

psi angles around this region. Well, if you go

back to that table, okay, alpha helix has a phi

angle of about minus 57 and psi angle of minus

47, but that's not an absolute value, okay? That's

sort of the average value. Depending on which

alpha helix, it's going to change by a few degrees

up and down, okay? And that's where you have,

okay, it's not exactly minus 47 and minus 57, but

it's going to be concentrated around those two values.

Left-handed alpha helices are very rare, but

they do exist, okay? And when you have left

-handed helices, you'll have phi-sci angles that

occupy this space in the Ramachandran plot.

All right, so we talked about the four secondary

structure elements, alpha helices, beta sheets,

beta turns, and random coils. Random coils, they

don't have a defined structure, so there's not a lot

to talk about. It's truly random. So, there's a very

nice equipment that most biochemistry departments

have. This is called a circular dichroism spectrometer,

often just called CD spectrometer. It's very

nice. You can insert your sample into this machine.

okay, it'll generate a plot depending on the

predominant type of secondary structure in your protein,

okay? So if your protein contains exclusively alpha

helices, you'll get a plot like this blue line.

If it's exclusively beta sheets, you'll get this

red profile. If it has both, like in this example,

it's going to be average value of those two

curves. So this is a technique that you can use to

determine which secondary structures are present

in your protein structure, okay?

All right, stay warm. I'll see you guys on Wednesday.