Protein Function|Graduate Biochemistry 4| Tulane

Protein Database Bank (PDB)

© Molview; PDBID=1bkv; Collagen

Structure and Function of Collagen

• extracellular proteins that provide rigidity and strength to many tissues
• Large bundles of triple helices
• Gly-X-Y

Alanine - Glycine substitute: A is hydrophobically contributed to α-helix composed.

Hydroxyproline…

• stabilized by cross-helix backbone hydrogen bonds
• large bundles of triple helices: A right-handed trimer of left-handed helices.
• Gly-X-Y Pattern: X is mostly proline and many of the Y is hydroxyproline (Gly-Pro-Hyp)

Glycine:

• No side chain: Closer the α-helix
• Hydrophobic: Folding to the center of the helix by hydrophobic effect

Alanine (replacing to Glycine):

• larger than Glycine but could still work similarly to Glycine

Prolein $ Hydroxyproline:

• pull out of the strand
• hydroxyproline work with water

Proline hydrolyzation was carried by Proline hydroxylase and required V_c as co-factor.

$$
Proline + 2-Oxoglutarate + O_ 2 + Fe^ {2+}\overset{Proline hydroxylase}{\longrightarrow} Hydroxyproline + Succinate + CO_ 2 + Fe^ {3+}
$$
$$
V_ c + Fe^ {3+} \to Dehydroascorbate + Fe^ {2+}
$$

• Collagen is stabilized by cross-helix backbone hydrogen bonds
  • Gly-NH::O=C-Pro

PS: Proline is often disturbing the structure of the α-helix. But why is proline’s favorite in the triple α-helix? (3-D structure)

Globin

Heme

• the oxygen binding cofactor
• Contains a reduced (ferrous, Fe²⁺) Iron atom.
• The porphyrin ring contains 4 pyrrole groups (A-D)
• Fe²⁺ has 6 coordination sites.
  • 4 pyrrole nitrogens
  • The proximal histidine that transmits ligand binding-induced conformational changes to the protein.
  • The sixth coordination site that binds various ligands O2, CO, CO2, CN
  • A “distal” histidine hydrogen-bonds to the heme-bound O2

Structure and function of myoglobin

• muscle protein that binds O2
• First protein to have its 3D structure determined
• contains a single heme group

© Molview; PDBID=1MBN; myoglobin

• First protein to have its 3D structure determined (by John Kendrew, 1956)
  • 153 residues long
  • 75% α-helical. No β-sheets.
  • There are 8 helices designated A-H

© Molview; PDBID=4hhb; hemoglobin

• α₂β₂ tetramer.
• Human: α, β, γ, δ
  • Consist to α₂X₂(X= β, γ or δ)
• β in adult; and γ in fetal.

© Cambridge University

Myoglobin (Mb) versus hemoglobin (Hb)

© Jay F Storz, 2002

• Hb: sigmoidal (26 torr).
• Mb: hyperbolic (2.8 torr).
• Mb has a greater affinity than Hb for oxygen at all oxygen pressures.
• In the lungs, Hb is saturated
• In tissues, oxygen is released from Hb and transferred to Mb .

© HarvardX

More information about Oxygen Transport: Karobben, Principal of Biochemistry

The Bohr (pH) Effect

Hemoglobin’s oxygen affinity decreases with decreasing pH and increasing CO2, causing the release of oxygen in tissues

PS: Bohr Effect is based on pH effect; pH Changes the conformation of the Myoglobin and Hemoglobin. (R state (oxy-form) to T state (deoxy-form))

• Protonatable His₁₄₆ of the β subunit is the key of Bohr effect.

• In the tissues (a low pH), His₁₄₆ forms a salt-bridge with Asp₉₄ and its backbone with Lys₄₀ of the, stabilizing the T state in tissues (Deoxy-form).
• In the lungs (a high pH), the salt-bridge is disrupted, promoting the T to R transition (Oxy-form).

H₁₄₆ of β interactions in T and R states

α₂Lys₄₀::β₁His₁₄₆::β₁Asp₉₄

© http://biomodel.uah.es

BPG effect:

© Molview; PDBID=1B86; BPG and Hemoglobin

• 2,3-bisphosphoglycerate (BPG) binds to hemoglobin (T state) and decreases oxygen affinity.
• BPG is produced within erythrocytes
• BPG is involved in high altitude adaptation
• BPG binds at the interface of β₁ and β₂ chains stabilized by numerous ionic and hydrogen bonding interactions
• Fetal hemoglobin (γ) doesn’t bind BPG
  • b/c His143 -> Ser mutation at the BPG binding site

© www3.nd.edu

Structure and Function of Antibody

• Antibodies function by interacting with foreign (antigen) molecules
• The antigen binding sites are produced by the variable domains from heavy and light chains.
• Several “hypervariable” loops are at the end of the variable domain.
• The two light chains and the two heavy chains are identical: the two antigen-binding sites are identical.

• Antibodies have H₂L₂ stoichiometry
  • H is a “heavy” chain and L is a “light” chain
• Each chain has one variable domain
• All domains (4 in each heavy chain and 2 in each light chains) have the same “immunoglobulin fold”
  • α β-sandwich with α 4-stranded and α 3-stranded sheet connected by a disulfide bond.

© nthu.edu

Antibody Peptide Interactions

• Strong and highly specific binding requires numerous favorable interactions between antibody and antigen.
  • Involving hydrophobic, ionic and hydrogen bonding interactions.

Video illustration

Myosin

© PDB; PDBID=1m8q; Myosin

• Sliding filament mechanism of muscle contraction:
  • During contraction, the thin and thick filaments move with respect to one another

• Two heavy chains and several light chains
• Heavy chains are rod-like molecules
  • The coiled coil tail domain
  • The ATPase head domain
• Arranged with tails overlapping and heads directed toward either end

Type	Images	Copyright	More
Actin		© PDB, ID=4pkg	- Long filamentous polymer consisting of two strands of globular monomers
Tropomyosin		© PDB, ID=1ic2	- A long, thin two-stranded α-helical rod - In relaxed muscle (low calcium), tropomyosin prevents myosin head from binding to actin
Troponin (three subunits)		© PDB, ID=1j1d	- Tn-I inhibits the actin-myosin interaction - Tn-T binds to tropomyosin - Tn-C binds calcium ions

Myosin Conformational Changes

© Takashi, et al.

Structural states of myosin during the contractile cycle.

1. Without bound nucleotide, myosin is strongly bound to actin (rigor state).
2. ATP binding to myosin dissociates the actin-myosin complex.
3. ATP is then hydrolyzed in ADP+Pi. There is a swing of the lever arm (green arrow).
4. Myosin can rebind to actin
5. Release its hydrolysis products and produce its force (green arrow) and again strongly bound to actin without nucleotide bound (power stroke, red arrow)