Proteins: General and spectroscopic properties Flashcards
All amino acids are chiral, with one exception. Which one?
Glycine (R-group = Hydrogen).
If a molecule is chiral, it can NOT be superimposed on its mirror image. Chiral centers are carbon atoms with four different ligands. The ligands positioning in space is what prohibits chiral reflections from being superimposed on one another.
Do naturally formed amino acids host a D- or L-chirality?
L-chirality (left-handed). This means that the “left mirror image” is the natural form of the amino acids.
Which are the positive polar amino acids?
Arginine
Lysine
Histidine (~10% of histidine is positive @ pH7)
Which are the negative polar amino acids?
Glutamate
Aspartate
What’s special about histidine with regards to polar charge?
Histidine can be positive or negative depending on the pH context.
What’s special about cysteine compared to other amino acids?
Two cysteins can form a disuflite bridge. This can occur within a protein or between two separate proteins.
What’s special with proline compared to other amino acids?
The R-group is a ring structure, which inhibits its ability to form hydrogen bonds, this makes proline a helix breaker.
What’s special about glycine?
- It’s not chiral (like all other amino acids)
- The R-group is hydrogen, this allows glycine to act like a hinge in helixes.
Explain what is displayed in the primary, secondary, tertiary and quarternary structures of a protein.
- Primary structure: Linear (displayed as a sequence)
- Secondary: local regularity (alfa helix, beta strands). Displayed linearly displayed as a sequence
2*. Super-secondary: local regularities (displayed in 2D) - Tertiary: The overall folding of a protein
4.Quarter: The interactions between multiple protein domains to make up the final protein complex.
Is the following statement true?: In alfa-helixes, the hydrogen groups are pointed outwards, the R-groups are pointed inwards.
No. The R-groups are pointed outwards. This makes sense as the R-groups are what facilitate the chemical reactions in which the protein can take place.
How many amino acid residues are there per turn in an alfa-helix?
3.6 resideus / turn
True or false: Are there parallel AND anti-parallel beta-sheets?
Yes.
What chemical interaction is instrinsic to disulfite bridges, zinc finger protein interactions and cytochrome interactions?
They are all covalent interactions.
Which two chemical interactions take part in the formation of salt bridges?
H-bonds, electrostatic bonds, ionic bonds
Why does base stacking improve the stability of a protein structure?
Base stacking generally occurs between residues which have planar ring structures, like tryptophan. The ring structures are generally hydrophobic, thus, they will be “attracted” to one another.
What are van der Waal interactions?
Short lived interactions between temporary dipole formations.
What are Edman degradation and Tandem mass spectrometry used for?
Edman degradation and tandem mass sspectrometry are used for assaying protein composition.
What are HPLC based analysis (glycosylation analysis) and mass spectronomy used for?
HPLC and mass spectronomy methods are used for assaying post-translational modifications. MS is used to find the mass / charge ratio, from which you can extrapolate the mass.
What are atomic absorption spectroscopy (AAS) and inductively coupled plasma mass spectrometry (ICP-MS) used for?
AAS and ICP-MS are used for metal ion analyses.
What does spectrofluorimetry measure?
The emitted light from the sample. Individual wavelengths are tested in order to characterize where absorption and emission occur.
Fluorescence is measured by a spectrometer. There is no detector behind the sample, thus, no spectrophotometer that observes transmitted light.
What does spectrophotometry measure?
The light absorbed by the sample.
What is the relationship between absorption and transmission?
Absorption: The amount of light which transfers energy to sample.
Transmission: The amount of light which passes through the sample.
Transmission = 1 - (absorption + scatter)
What’s the relationship between luminescence, fluorescence and phosphorescence?
Fluorescence + phosphorescence = Luminescence.
Phosphorescence remains when the light source is removed. Fluorescence only refers to the immediate light reaction after having excited a chromophore.
What’s the difference between a spectrophotometer and a spectrometer?
A spectrometer has its detector 90 degrees off-angle from the illumination source. Transmitted light is not observed. Detects sample luminescence.
A spectrophotometer has its measurement device straight behin the sample, observing transmittance.
what’s “quantum yield” a measurement of?
it measures the efficiency of fluorescence.
Photons emitted from sample / samples absorbed by the sample.
What three mechanims can cause fluorophore quenching?
Dynamic queenching (via collisions): The fluorophore is deactivated through interactions with other molecules.
Static quenching: The fluorophore is statically bound in a complex.
Resonance energy transfer: The fluorophore absorbs light, gets excited, emits a wavelength, which in turn is absorbed by another fluorophore, that fluoresces.
Name an application of fluorophore quenchning.
Assaying whether a protein is folded or not. The less folded the protein, the more fluorescence.
What’s is FRET? what’s the mechanism of FRET? Give an example of two fluorophores which have this relationship.
FRET = Fluorescence resonance energy transfer.
Two fluorophores are near each other, one of them is excited, which in turn excites the other one. CFP and YFP have this relationship.
FRET can be used to see if two proteins are in interactive range by ex fusing CFP and YFP to different proteins and assaying whether FRET can occur.
