Protein Folding & Biospectroscopy Lecture 6 F14PFB David Robinson.
Protein Folding & Biospectroscopy F14PFB Dr David Robinson Lecture 2.
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Transcript of Protein Folding & Biospectroscopy F14PFB Dr David Robinson Lecture 2.
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Protein Folding & Biospectroscopy
F14PFB
Dr David Robinson
Lecture 2
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Principles of protein structure and function
• Function is derived from structure
• Structure is derived from amino acid sequence
• Different activities and shapes of proteins due to different amino acid sequences
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A reminder…• Basic Amino Acid
Structure:– The side chain, R,
varies for each ofthe 20 amino acids
C
RR
C
H
NO
OHH
H
Aminogroup
Carboxylgroup
Side chain
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The Peptide Bond
• Dehydration synthesis
• Repeating backbone: N–C –C –N–C –C
– Convention – start at amino terminus and proceed to carboxy terminus
O O
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Levels of Protein StructureLevels of Protein Structure
- helix
Myoglobin
Hemoglobin
The folded protein structure is stabilized by a variety of weak chemical interaction, and in some cases covalent (disulfide) bonds between cysteine residues
R– CH2–S–S–CH2–R
Cys Cys
Disulfide bond:
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Structural element Description
primary structure amino acid sequence of protein
secondary structure helices, sheets, turns/loops
super-secondary structure association of secondary structures
domain self-contained structural unit
tertiary structure folded structure of whole protein
• includes disulfide bonds
quaternary structure assembled complex (oligomer)
• homo-oligomeric (1 protein type)
• hetero-oligomeric (>1 type)
Protein structure:Protein structure: overview overview
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Primary & Secondary StructurePrimary & Secondary Structure Primary structurePrimary structure = the linear sequence of
amino acids comprising a protein:AGVGTVPMTAYGNDIQYYGQVT…
Secondary structureSecondary structure• Regular patterns of hydrogen bonding in proteins
result in two patterns that emerge in nearly every protein structure known: the -helix and the-sheet
• The location of direction of these periodic, repeating structures is known as the secondary secondary structurestructure of the protein
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The alpha helixThe alpha helix 60°
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Properties of the alpha helixProperties of the alpha helix 60° Hydrogen bondsHydrogen bonds
between C=O ofresidue n, andNH of residuen+4
3.6 residues/turn 1.5 Å/residue rise 100°/residue turn
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Properties of Properties of -helices-helices 4 – 40+ residues in length Often amphipathic or “dual-natured”
• Half hydrophobic and half hydrophilic• Mostly when surface-exposed
If we examine many -helices,we find trends…• Helix formers: Ala, Glu, Leu,
Met• Helix breakers: Pro, Gly, Tyr,
Ser
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The beta strand (& sheet)The beta strand (& sheet) 135° +135°
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Properties of beta sheetsProperties of beta sheets Formed of stretches of 5-10 residues in
extended conformation Pleated – each C a bit
above or below the previous Parallel/antiparallelParallel/antiparallel,
contiguous/non-contiguous
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Parallel and anti-parallel Parallel and anti-parallel -sheets-sheets Anti-parallel is slightly energetically favoured
Anti-parallelAnti-parallel ParallelParallel
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Turns and LoopsTurns and Loops Secondary structure elements are connected by
regions of turns and loops Turns – short regions
of non-, non-conformation
Loops – larger stretches with no secondary structure. Often disordered.• “Random coil”• Sequences vary much more than secondary
structure regions
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Levels of Protein Levels of Protein StructureStructure
Secondary structure elements combine to form tertiary structure
Quaternary structure occurs in multienzyme complexes• Many proteins are
active only as homodimers, homotetramers, etc.
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Protein Folding• Forming polypeptide chain requires energy
and information (template) – ie translation from RNA protein SEQUENCE
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Protein Folding• Forming polypeptide sequence requires
energy and information (template)
• Forming native conformation requires NO ADDITIONAL energy or information
(SELF ASSEMBLY)
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Protein folding
Amino acid sequence contains all information necessary for folding into a specific three-dimensional structure
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Protein Folding
Proteins, in general, do NOT fold as they are synthesized on the ribosome
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Folding of RNAse A in the test tubeFolding of RNAse A in the test tube
denaturation renaturation
Incubate proteinin guanidine
hydrochloride(GuHCl)or urea
100-folddilution of proteininto physiological
buffer
Anfinsen, CB (1973) Principles that govern the folding of protein chains. Science 181, 223-230.
- the amino acid sequence of a polypeptide is sufficient to specify its three-dimensional conformation
Thus: “protein folding is a spontaneous process that does not require the assistance of extraneous factors”
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Protein Folding
Many proteins fold by Assisted Self Assembly
Correct assembly (native conformation) requires assistance
by CHAPERONES
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Protein unfolding = DenaturationLoss of structure and function
– Heat– Extreme pH– Detergents– Urea
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Protein unfolding = DenaturationWhy do these conditions cause loss of
structure and function?– Heat– Extreme pH– Detergents– Urea
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Lysozyme
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Lysozyme
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Tertiary: complete three-dimensional
structure
Quaternary: arrangement of subunits
(in multisubunit protein)
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Hemoglobin
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Quaternary structure
• Held together by weak interactions between side (R/functional) groups as well as covalent disulfide bonds
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Structure-function relationship• Function is derived from structure
• Structure is derived from sequence
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Sickle-cell diseaseNormal red blood cells Sickle shaped red blood cells
Due to single amino acid change in haemoglobin
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Sickle-cell disease
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Sickle-cell disease
• Single specific amino acid change causes change in protein structure and solubility
• Results in change in cell shape
• Causes cells to clog blood vessels
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Amino acids