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Fundamentals of protein and
nucleic acid structure
Lecture 2
Structural Bioinformatics
Dr. Avraham Samson
81-871
Tree of Life
All known life forms use the same building blocks suggesting there was a common ancestor
The tree of life is not that simple!
Gene transfer across kingdoms continually occurs
(cyanobacteria became chlorophylls, proteobacteria mitochondria, viruses)
Inside living cells
29 atoms of life
Most common elements: H, O, C, N, P, S (97% of organism weight)
Most common ions: Ca2+, K+, Na+, Mg2+, Cl-
Forces affecting structure:
•
•
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H-bonding
Van der Waals
Electrostatics
Hydrophobicity
Disulfide Bridges
d
2.6 Å < d < 3.1Å
150° < θ < 180°
Forces affecting structure:
•
•
•
•
•
H-bonding
Van der Waals
Electrostatics
Hydrophobicity
Disulfide Bridges
Repulsion
‫דחייה‬
Attraction
‫משיכה‬
d
3 Å < d < 4Å
Forces affecting structure:
•
•
•
•
•
H-bonding
Van der Waals
Electrostatics
Hydrophobicity
Disulfide Bridges
Coulomb’s law
d
d = 2.8 Å
“IONIC BOND” ‫קשר יוני‬
“SALT BRIDGE” ‫גשר מלח‬
E = Energy
k = constant
D = Dielectric constant (vacuum = 1; H2O = 80)
q1 & q2 = electronic charges (Coulombs)
r = distance (Å)
Forces affecting structure:
•
•
•
•
•
H-bonding
Van der Waals
Electrostatics
Hydrophobicity
Disulfide Bridges
Forces affecting structure:
•
•
•
•
•
H-bonding
Van der Waals
Electrostatics
Hydrophobicity
Disulfide Bridges
Other names:
cystine bridge
disulfide bridge
Hair contains lots of disulfide bonds
which are broken and reformed by heat10
Levels of protein structure
Primary: Amino acid sequence
Secondary: Local fold pattern of small subsequence
Tertiary: Fold of entire protein chain
Quaternary: Complex of multiple chains
Primary (1o) structure
The amino acid sequence is the primary structure
Primary (1o) structure
20 amino acids
13
Primary (1o) structure
14
Primary (1o) structure
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pKa = pH of 50%
dissociation
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Primary (1o) structure
Amino acid nomenclature
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Primary (1o) structure
• Amino acids polymerize through peptide
bonds to form polypeptides
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Primary (1o) structure
• N-terminal is the start of a polypeptide chain
• Amino acids are also called residues
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Primary (1o) structure
side chains ‫שייר צדדי‬
backbone ‫שלד‬
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Primary (1o) structure
• Post translational modification are important
because they can change the function of proteins
(i.e. phosphorylation, acetylation, hydroxylation,
carbohydrate and lipid modifications)
N-terminal acetylation
O-phosphotyrosine
hydroxyproline
γ-carboxyglutamate
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Amino acids chirality
Enantiomers –mirror images - ‫תמונת ראי‬- ‫אננטיומרים‬
chiral center C ‫מרכז כיראלי‬
Dextro-Laevus in Latin
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‫‪Most proteins: only L amino acids‬‬
‫‪C chiral center is L configuration‬‬
‫איך לקבוע קונפיגורציה ‪ L‬ו‪D-‬‬
‫•סדר את האטומים לפי עדיפות על פי‬
‫הכללים הבאים‬
‫כלל ‪ .1‬אטום עם מספר אטומי יותר גבוה‬
‫בעל סדר עדיפות יותר גבוה‪.‬‬
‫)‪(I > Cl > O > N> C > H‬‬
‫כלל ‪ .2‬אם האטומים זהים‪ ,‬העדיפות על‬
‫פי האטומים המתמירים‬
‫)‪(C(CH3)3 > CH(CH3)2 > CH2CH3 > CH3‬‬
‫•שים את האטום עם סדר העדיפות הכי‬
‫נמוך מאחור‪.‬‬
‫• קבע את כיוון החץ ‪ ,‬מסדר עדיפות הכי‬
‫גבוה לככוון הכי נמוך ‪.‬אם החץ‪:‬‬
‫‪ 23‬רמז יותר קל בשביל לזכור ‪) CORN :‬תירס(‬
‫עם כיוון השעון קונפיגורציה ‪D‬‬
‫נגד כיוון השעון קונפיגורציה ‪L‬‬
Bond angles
Torsion angles w, φ, and ψ
• Unlike w, the two backbone dihedral angles φ
and ψ are free to rotate
• This rotation freedom allows protein folding
w
Dihedral angles:
-180o < φ < +180o
-180o < ψ < +180o
w is 0o or 180o
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Peptide bond is planar
Cα, C, O, N, H, Cα all lie in the
same plane
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Torsion angle (w) is usually trans
Steric hindrance
Question: What other residues can be cis?
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Except for X-Pro bond in which cis is
preferred
Steric hindrance
Steric hindrance
Steric hindrance allows both cis and trans (4:1 ratio)
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Ramachandran Diagrams
Preferred regions
• Steric hindrance dictates torsion angle preference
• Ramachandran plot show preferred regions of φ and ψ
dihedral angles
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Secondary (2o) structure
Helices
• -helix
• 310-helix
• p-helix
b Sheets
• Antiparallel
• Parallel
Turns
• b,g,,and p
Unstructured
α-helix is the most
common
3.6 residues per turn
(number of residues in
one full rotation of
360°)
5.4 Å pitch (translation
along axis for one full
rotation of 360°)
Hydrogen bond: i→i+4
Secondary (2o) structure
Helices
• -helix
• 310-helix
• p-helix
b Sheets
• Antiparallel
• Parallel
Turns
• b,g,,and p
Unstructured
310-helices are
rare in proteins:
3.1 residues per
turn (number of
residues in one full
rotation of 360°)
6 Å pitch
(translation along
axis for one full
rotation of 360°)
Hydrogen bond: i→i+3
Secondary (2o) structure
Helices
• -helix
• 310-helix
• p-helix
b Sheets
• Antiparallel
• Parallel
Turns
• b,g,,and p
Unstructured
π -helices are
rare in proteins
4.3 residues per
turn (number of
residues in one full
rotation of 360°)
6 Å pitch
(translation along
axis for one full
rotation of 360°)
Hydrogen bond: i→i+5
Hydrogen bonding in helices
• CO of residue (n) forms an h-bond with NH of residue:
– (n+4) → α-helix
– (n+3) → 310-helix
– (n+5) → π-helix
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Ribbon diagram of α-helix
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Ramachandran of α-helix
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Secondary (2o) structure
Helices
• -helix
• 310-helix
• p-helix
b Sheets
• Antiparallel
• Parallel
Turns
• b,g,,and p
Unstructured
In antiparallel b-sheets
•Adjacent β-strands run in opposite directions
•Hydrogen bonds (dashed lines) between NH
and CO stabilize the structure
•The side chains (in green) are above and
below the sheet
Secondary (2o) structure
Helices
• -helix
• 310-helix
• p-helix
b Sheets
• Antiparallel
• Parallel
Turns
• b,g,,and p
Unstructured
In parallel b-sheets
•Adjacent β-strands run in same direction
•Hydrogen bonds (dashed lines) between NH
and CO stabilize the structure
•The side chains (in green) are above and
below the sheet
Ribbon diagram of β sheet
• In addition to being purely parallel or antiparallel, β
sheets can be mixed, with strands running in both
parallel and antiparallel directions
• Arrow pointing to C-terminal end
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Ramachandran of β-sheet
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Secondary (2o) structure
Helices
• -helix
• 310-helix
• p-helix
b Sheets
• Antiparallel
• Parallel
Turns
• b,g,,and p
Unstructured
CO of residue i forms
h-bonds with NH of
either residue i+2,
i+3, i+5, or i+4
β-turn i→i+3 H-bond (most
common)
γ-turn: i→i+2 H-bond
α-turn i→i+4 H-bond
π-turn i→i+5 H-bond
Secondary (2o) structure
Helices
• -helix
• 310-helix
• p-helix
b Sheets
• Antiparallel
• Parallel
Turns
• b,g,,and p
Unstructured
Tertiary (3o) structure
• Proteins fold into compact tertiary structures with
hydrophobic cores. (spacefill representation)
43
Structural classification: http://www.cathdb.info/cgi-bin/cath/GotoCath.pl?link=class.html
Tertiary (3o) structure
• It is a bit difficult to see and understand
anything here (sticks representation)
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Tertiary (3o) structure
• To simplify representation, secondary
structure diagrams are used
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Quaternary (4o) structure
Dimer of identical subunits (Cro protein of bacteriophage lambda)
Quaternary structure
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Coat of rhinovirus
(common cold)
60 copies of 4
different subunits,
3 outside,
red, blue, green
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Protein structure databases
• Primary: UniProt http://www.uniprot.org/
• Secondary: DSSP http://rcsb.org
• Tertiary: PDB http://rcsb.org
• Quaternary: PQS http://www.ebi.ac.uk/pdbe/pqs/
DNA and
RNA Structure
NOTE:
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• Components
• Sugar
• Base
• Phosphate
5’ to 3’ direction
T->U in RNA
RNA - extra –OH at 2’ of
pentose sugar
DNA - deoxyribose
Numbering
• Single vs double
strands
• DNA more stable
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p. 792.
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The 5 Bases
of DNA and RNA
Purines
NOTE:
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Pyrimadines and Purines
T->U in RNA
Names
Numbering
Bonding character
Position of hydrogen
Tautomers
Pyrimadines
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Tautomeric
Structures
• Keto vs enol (OH)
• Different hydrogen
bonding patterns
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Geometry of
Watson Crick
Base Pairs
• A:T and G:C pairs are
spatially similar
• 3 H-bonds vs 2 (GC rich?)
• Sugar groups are attached
asymmetrically on the same
side of the pair
• Leads to a major and
minor grove
• Bases are flat but the
hydrogen bonding leads to
considerable flexibility
• Base stacking is flexible
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p. 797.
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Definition of Major
and Minor Groove
Hydrogen bonding
of WC base pair
Mechanisms of
recognition
The canonical Watson-Crick base pair, shown as the G-C pair. Positions of the
minor and major grooves are indicated. The glycosidic sugar-base bond is shown
by the bold line; hydrogen bonding between the two bases is shown in dashed
lines.
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Base
Morphology
The base-pair reference frame is
constructed such that the x-axis
points away from the (shaded)
minor groove edge.
Images illustrate positive values
of the designated parameters.
Reprinted with permission from
Adenine Press from (Lu, et al., 1999).
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Backbone
Conformation
Voet, Donald and Judith G. Biochemistry.
John Wiley & Sons, 1990, p. 807.
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A Beta-nucleoside
• Ring is never flat – has 5
internal torsional angles
• The pucker is determined
by what is bound
• A variety of puckers have
been observed
• Pucker has a strong
influence on the overall
conformation
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The Glycosidic Bond
Anti
Syn
• Connects ribose sugar to the base
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Change in sugar
conformation
affects the
backbone
C3’
C2’
C3’-Endo
C3’
C2’
C2’-Endo
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..and the
position of A DNA
the bases
relative to
the helix axis
B DNA
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Canonical B DNA
Neidle, Stephen. Nucleic Acid Structure and Recognition.
Oxford University Press, 2002, p. 34.
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Canonical B DNA
• First determined experimentally by fiber diffraction
(Arnott)
• C2’-endo sugar puckers
• High anti glycosidic angles
• Right handed – 10 base pairs per turn
• Bases perpendicular to the helix axis and stacked over the
axis
• Overall bending as much as 15 degrees (result of base
morphologies – twist and roll) – {machine learning –
sequence vs overall conformation?}
• Over 230 structures 25 with base mis-pairing – only cause
local perturbations
• Strong influence of hydration along spine
http://ndbserver.rutgers.edu/index.html
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A DNA
Neidle, Stephen. Nucleic Acid Structure and Recognition.
Oxford University Press, 2002, p. 36.
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Canonical A DNA
• C3’-endo sugar puckers – brings consecutive
phosphates closer together 5.9A rather than 7.0
• Glycosidic angle from high anti to anti
• Base pairs twisted and nearly 5A from helix axis
• Helix rise 2.56A rather than 3.4A
• Helix wider and 11 base pairs per repeat
• Major groove now deep and narrow
• Minor grove wide and very shallow
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Z-DNA
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Z-DNA
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Helix has left-handed sense
Can be formed in vivo, given proper sequence and superhelical tension, but
function remains obscure.
Narrower, more elongated helix than A or B.
Major "groove" not really groove
Narrow minor groove
Conformation favored by high salt concentrations, some base substitutions,
but requires alternating purine-pyrimidine sequence.
N2-amino of G H-bonds to 5' PO: explains slow exchange of proton, need for
G purine.
Base pairs nearly perpendicular to helix axis
GpC repeat, not single base-pair
– P-P distances: vary for GpC and CpG
– GpC stack: good base overlap
– CpG: less overlap.
•
•
Zigzag backbone due to C sugar conformation compensating for G glycosidic
bond conformation
Conformations:
– G; syn, C2'-endo
– C; anti, C3'-endo
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Z-DNA
•
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•
Convex major groove
Deep minor groove
Alternate C then G
Spine of hydration
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Quadruplex DNA
1NP9
Jmol
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tRNA
Invariant L-shape
1EVV
jmol
Saenger, Wolfram. Principles of Nucleic Acid Structure.
Springer-Verlag New York Inc., 1984, p. 333.
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tRNA H
bonds
between
distant
regions
Neidle, Stephen. Nucleic Acid Structure and Recognition.
Oxford University Press, 2002, p. 148.
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The Ribosome
• Complex of protein
and RNA
• Small 30S subunit –
controls interactions
between mRNA and
tRNA
• Large 50S subunit –
peptide transfer and
formation of the
peptide bond
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Nucleic acid structure databases
• Primary: UniProt http://www.uniprot.org/
• Tertiary: PDB http://rcsb.org contains all of
http://ndbserver.rutgers.edu/
Molecular viewers
• Pymol
• Rasmol
• Jmol
• Molmol
• VMD
• Swiss PDB
etc…
(please note, only freeware are listed)
Transcription and translation (DNA  RNA  Protein)
Transcription and translation (DNA  Protein)