Quiz L3(end stuff) - L8

Role of Hemoglobin(Hb) vs Myoglobin(Mb)

Hemoglobin - Transport oxygen throughout our bloodstream + deliver it to all tissues in our body 

Myoglobin - To store oxygen in muscle tissues + release to different tissues when oxygen is needed. 

How many subunits do Hemoglobin vs Myoglobin have

Hemoglobin = 4 subunits so can bind 4 oxygen molecule

Myoglobin = 1 subunits so can bind 1 oxygen molecule

What organelle does Mb send O₂ to and what is O₂ consumed for

Mitochondria for the ETC

Formation of MbO₂ increase/decrease the total amount of O₂ in the cytosol by 300-fold

increases

What is the heme group in Mb

Fe²⁺ (only form that can be used to accept oxygen)

How many coordination is the Fe²⁺ in Mb and how many are ocupied by nitrogen

6 coordinations for Fe²⁺

4 occupied by nitrogen

5th group occupied by proximal HIS(has a nitrogen from his) which holds the heme in the F helix Mb

6th group occupied by distal HIS which allows binding to O2 in the heme pocket of the E helix of Mb

Without Mb, Water binds to Fe²⁺ hemewhich causes what

Fe²⁺ heme will become Fe³⁺ heme

• Oxygen reacts with heme but doesnt bind with heme

What is the function of Mb polypeptide

• Keep the heme Fe2+ close to the water, where oxygen is;(hydrophilic on exterior and heme on the hydrophobic interior

• Keep the heme Fe2+ away from contact with all other hemes, because it is actually stacks of hemes that chemically react with the oxygen.

• Enable hundreds of times higher concentration of oxygen than could be achieved in water, since Mb is so soluble in the water inside a cell and free O2 is not.

The reason that muscle is red is due to

Myoglobin

Is the Mb binding of Oxygen a reversible process

Yes since no covalent bond is formed so the reaction can run right to left but its favored the right

What is a Prosthetic group, Holoprotein and Apoprotein

Prosthetic Group - Non AA(heme in the case of Mb) structure covalently bonded in the protein

Holoprotein - entire protein + prossthetic group

Apoprotein - protein w/o its prosthetic group

What is Hemoglobin and Myoglobin a Holoprotein vs Apoprotein

Holoprotien

Hb = binded to heme

Mb = binded to O2

Apoprotein

Hb = not binded to heme

Mb = not binded to O2

How many methods are there to find amino acid sequence(primary structure) and what are they

2 methods

1. Protien Based(older method via edman degradation) - each amino acid is cleaved and identified

2. DNA Based(newer method) - obtain a small bit of the AA sequence via method 1(i.e mass spec) and will identify the sequence into nucleotides

What are Isozymes = isoenzyme = isoforms

enzymes that are structurally(AA sequence & 3D fold) and catalytically similar but not identical

i.e hexokinase

Why do we have different Hexokinase in different cell types

HK IV binds to glucose weakly but when glucose conc is high then HK IV does bind glucose and catalyze rxn to form glucose phosphate

What are Homologs and the 2 types

Homologs = shared common ancestor

• Paralogs = in same speices

• Orthologs = in different speices

What can AA sequence do if not 3D structure yet

1) Function: enzyme activity

• Comparison of an unknown AA sequence with a known sequence(similar sequences)

2) Function: Building Domains

• Proteins or parts of proteins that are not catalysts (so no chemical reactions)

What are the 2 Evolutrionary Relationship of AA sequences

1) Sequence of the same protein from different organisms(that are quite different) can lead to a phylogenetic tree

• i.e cytochrome C is conserved

2) Find Special short sequences

• little signals that are found in nature, like how serine gets phosphorylated by a kinase or telling a protein to go to certain location

What is AlphaFold 2 (by deepmind,london, UK: Computational Method)

Predicts 3D protein structure given the AA sequence

What is Bioinformatics

In biology there is a vast amount of information and these info need special treatment

What are the 3 main ways atomic 3D structure protein can be determined(Experimental method)

1) X-Ray Diffraction of protein CRYSTALS

• 85% of protein found this way

• Hardest part is to grow the crystal

2) Multi-Dimensional NMR of protein dissolved in buffer

• 15% of protein found this way

• requires larger amount of proteins

3) Cryo Electron Microscopy (atomic level at 4 A)

• yields atmoic level

Who is Dorothy Crowfoot Hodgkin & Kendrew & Perutz

Hodgkin: 1964 Nobel Prize Winner for proving how x-ray crystallography could directly map the atomic positions of complex biological molecules

Kendrew & Perutz: Nearly all amino acid positions located in 3D for myoglobin

Describe Light

• Packs of photons

• Wave and particle properties through X-Ray beam (high energy light) with an oscillating electromagnetic wave

• Range of X-Ray wavelengths possible 

What is the Phase problem in X-Ray Crystallography

X-rays are shot at protein crystals, leading to scattering in all directions by every atom. However, during the process, only information about the intensity of the light was found, which yielded missing information on the phase of the light.

It took them 30 years to find the phase information, suggesting that formation of good crystals was a limiting factor.

What are the 4 ways to show protein strucutre

Sticks

Spheres

Ribbon

Cartoon

What are the 6 categories of non-covalent bonds important for constrruction of protein structure

1) Hydrogen Bonds - H with FON

2) Electrostatic - every positive and negative must have an equal opposite charge

3) Disulfide - weak for covalent but stronger than H-Bond

4) Weak and weakly directional forces - dipoles, London dispersion

5) Ring Stacking (Phe, Tyr & Trp)

6) Hydrophobic interactions - favorable water entropy

What does it mean that a protein crystal is 50% water

Salts when in crystal have no empty space whereas proteins have empty spaced filled by fluid

What does it mean that alpha helix(8 alpha helices) 70% Mb

Mb has one of the higest alpha helix content and some protein have no alpha helix

What does it mean by Heme in a crevice for Mb

Protein makes precise fit , Fe²⁺ kept away from water

What are the 6 atoms in the peptide plan

1) C_alpha of residue i

2) C (carbonyl carbon) of residue i

3) O (carbonyl carbon) of residue i

4) N (amide nitrogen) of residue i+1

5) H (amide hydrogen) of residue i+1

6) C_alpha of residue i+1

Why are the 6 atoms in the peptide plane constrained in a plane

Resonance between C-O and C-N, each having partial double bond character.

What is the Boltzmann Distribution equation and what does it tell you

Boltzmann Distribution equation: P = e^-x/RT

P = the probability of the certain energy cost being sufficient to reach a certain deviation from the ground state

x = the amount of energy needed to reach a certain deviation from the ground state (in kJ/mol)

R = the gas constant which is equal to 8.31 J/mol * K

T = the temperature (in Kelvin)

Equation tells you the conformation of known energy relative to ground state

What are the bonds corresponding to backbone torsion angles phi, psi and omega

Torrison Angle or Phi (ɸ) → bond between Cɑ and N → 180 degrees

Psi (Ѱ) → bond between Cɑ and C → 180 degrees

Omega (Ѡ) → bond between C and N (peptide bond) → 0 degrees

Explain the info displayed on a Ramachandran Map

The info shows the possible combination of torsion angle values (ɸ and Ѱ) in a secondary structure of polypeptide due to steric hindrance

Each point on the plot shows a single amino acid’s torsion angles, rather than the entire protein

We cannot tell the 3D structure information of protein from the graph as we are missing the primary amino acid sequence information.

Note: White region are allowed region = no steric hinderance

Where are the general locations of the alpha helix and beta sheets

What is the meaning of calculated vs.“real protein” Ramachandran Plot

Calculated Ramachandran Plot provides an estimate for the allowed regions (theoretical modeling based on steric constraints) while “real protein” Ramachandran Plot provides the actual, empirical database plotting of ɸ, Ѱ value combinations for a few 100 proteins.

Calculated: shaded region is forbidden

Real data: shaded region is where you find the protein

Explain why the Ramachandran Plots for hundreds of folded and unfolded proteins look similar

Both folded and unfolded proteins are subject to same physical constraint of peptide backbone, so certain combinations of ɸ and Ѱ torsion angles are energetically unfavorable and not always sterically possible.

Alpha helix, beta sheet, turns, collagen are the four secondary structures found -> polyproline type II helix