BCB 444544 Introduction to Bioinformatics

1
BCB 444/544 - Introduction to Bioinformatics
Lecture 29 Protein Structure Basics
Prediction 29_Nov1
2
Seminars in Bioinformatics/Genomics

• Mon Oct 30
• Madan Bhattacharyya (Agron, ISU) Isolation of
  Signaling Genes for Phytophthora Resistance in
  Soybean
• IG Faculty Seminar 1210 PM in 101 Ind Ed II
• Thurs Nov 2
• Peter Clote (ComS, Boston Univ) Some New
  Aspects of Protein
• RNA Structure
• Baker Center Seminar 210 PM in Howe Hall
  Auditorium
• Laura Dutca (Chem, ISU) Detailed analysis of
  E.coli primary r-Protein Interaction with 16rRNA
  Implications for RNP assembly
• BBMB Seminar 410 PM in 1414 MBB
• Fri Nov 3
• Heather Greenlee (BMS, ISU) Decoding the
  Rod-Photoreceptor Differentiation Pathway
• BCB Faculty Seminar 210 PM in W142 Lago
• Eric Henderson (GDCB, ISU) Putting the "Bio" in
  Bionanotechnology
• GDCB Seminar 410 PM in 1414 MBB

3
Assignments Reading This Week

• Chp 8 Proteomics
• More Machine Learning Algorithms
• vMon Oct 30 Chp 8.1 Proteomics - Introduction
  Chp 8.2 Protein 3D Structure
• vWed Nov 1 Chp 8.3 Protein Interaction Networks
• Chp 8.4 Measuring Proteins
• Thurs Nov 2 Lab 9 Attendance Seminar
  Feedback Form (immediately after seminar)
  required
• Peter Clote 210 PM Howe Hall Auditorium New
  Aspects of Protein RNA Structure
• Fri Nov 3 Machine Learning - more Algorithms
• Support Vector Machines (SVMs)
• Neural Networks (ANNs)

4
Assignments Events
Exam 2 Keys Grades posted today Exam returned
today after class BCB 444 544 HW5
Posted online yesterday (sorry) Due by Noon,
Mon Nov 6 BCB 544 Only Teams Projects
Any questions? 544Extra2 Due Mon
Nov 6
See updated Schedule (Oct 30) posted online
5
Protein Structure Function

• Protein Structure
• Amino acids characteristics
• Structural classes motifs
• Protein functional families
• Classification
• Databases
• Visualization

6
Protein Structure Function

• Protein structure - primarily determined by
  sequence
• Protein function - primarily determined by
  structure
• ( structure determines interactions with other
  molecules)
• Globular proteins
• have compact hydrophobic core hydrophilic
  surface
• Membrane proteins
• have special hydrophobic domains
• often transmembrane (TM) helices

7
Protein Structure Function

• Protein Folding?
• Folded proteins are only marginally stable,
  because proteins must balance stability vs
  function
• Intrinsically disordered some domains of
  proteins (or even entire proteins) that do not
  assume a stable "fold" until they are bound to
  their partner (protein, DNA, etc.)
• Predicting protein structure and function can be
  very difficult -- but is increasingly important

8
4 Basic Levels of Protein Structure
9
Amino Acids

• Each of 20 different amino acids has different
  "R-Group" side chain attached to Ca

10
Peptide bond is rigid and planar
11
Hydrophobic Amino Acids
12
Charged Amino Acids
13
Polar Amino Acids
14
Certain side-chain configurations are
energetically favored (rotamers)
Ramachandran plot "Allowable" psi phi angles
15
Glycine is smallest amino acidR group H atom

• Glycine residues increase backbone flexibility
  because they have no R group

16
Proline is cyclic

• Proline residues reduce flexibility of
  polypeptide chain
• Proline cis-trans isomerization is often a
  rate-limiting step in protein folding
• Recent work suggests it also regulates ligand
  binding in native proteins -Andreotti

17
Cysteines can form disulfide bonds

• Disulfide bonds (covalent) stabilize
• 3-D structures
• In eukaryotes, disulfide bonds are found only in
  secreted proteins or extracellular domains

18
Globular proteins have a compact hydrophobic core

• Packing of hydrophobic side chains into interior
  is main driving force for folding
• Problem? Polypeptide backbone is highly polar
  (hydrophilic) due to polar -NH and CO in each
  peptide unit these polar groups must be
  neutralized
• Solution? Form regular secondary structures,
• e.g., ?-helix, b-sheet, stabilized by H-bonds

19
Exterior surface of globular proteins is
generally hydrophilic

• Hydrophobic core formed by packed secondary
  structural elements provides compact, stable core
• "Functional groups" of protein are attached to
  this framework exterior has more flexible
  regions (loops) and polar/charged residues
• Hydrophobic "patches" on protein surface are
  often involved in protein-protein interactions

20
Secondary Structural Elements

• ??Helix
• ?? Sheets
• Loops
• Coils

21
?a?- Helix

• Most abundant 2' structure in proteins
• Average length 10 aa's (10 Angstroms)
• Length varies from 5-40 aa's
• Alignment of H-bonds creates dipole moment
  (positive charge at NH end)
• Often at surface of core, with hydrophobic
  residues on inner-facing side, hydrophilic on
  other side

22
??helix is stabilized by H-bonds between every
4th residue
C black O red N blue
23
R-groups are on outside of ??helix
24
Types of ??helices

• "Standard" ??helix 3.6 residues per turn
• H-bonds between C0 of residue n and
• NH of residue n 4
• Helix ends are polar almost always on surface of
  protein
• Other types of helices?
• n 5 ? helix
• n 3 310 helix

25
Certain amino acids are "preferred" others are
rare in ??helices

• Ala, Glu, Leu, Met good helix formers
• Pro, Gly, Tyr, Ser poor
• Amino acid composition distribution varies,
  depending on location of helix in 3-D structure

26
??-Strands Sheets

• H-bonds formed between 5-10 consecutive residues
  in one portion of chain with another
• set of 5-10 residues farther down chain
• Interacting regions may be adjacent (with short
  loop between) or far apart
• ?-sheets usually have all strands either parallel
  or antiparallel

27
Antiparallel???-sheet
28
Antiparallel???-sheet
29
Parallel???-sheet
30
Mixed??-Sheets also occur
31
?Loops

• Connect helices and sheets
• Vary in length and 3-D configurations
• Are located on surface of structure
• Are more "tolerant" of mutations
• Are more flexible and can adopt multiple
  conformations
• Tend to have charged and polar amino acids
• Are frequently components of active sites
• Some fall into distinct structural families
• (e.g., hairpin loops, reverse turns)

32
Coils

• Regions of 2' structure that are not helices,
  sheets, or recognizable turns
• Intrinsically disordered regions appear to play
  important functional roles

33
Globular proteins are built from recurring
structural patterns

• Motif or supersecondary structure
• combination of 2' structural elements
• Domain combination of motifs
• Independently folding unit (foldon)
• Functional unit

34
A few common structural motifs

• Helix-turn-helix e.g., DNA binding
• Helix-loop-helix e.g., Calcium binding
• ?b-hairpin 2 adjacent antiparallel strands
• connected by short loop
• Greek key 4 adjacent antiparallel strands
• b?a-b 2 parallel strands connected by helix

35
H-T-H H-L-H
36
?b-hairpin
37
Greek key
38
Beta-alpha-beta
39
Simple motifs combine to form domains
40
Large polypeptide chains fold into several domains
41
6 main classes of protein structure

• 1) a Domains
• Bundles of helices connected by loops
• 2) ? Domains
• Mainly antiparallel sheets, usually with 2 sheets
  forming sandwich
• 3) a????Domains
• Mainly parallel sheets with intervening helices,
  also mixed sheets
• 4) ??? a????Domains
• Mainly segregated helices and sheets
• 5) Multidomain ?????????
• Containing domains from more than one class
• 6) Membrane cell-surface proteins

42
?-domain structures coiled-coils
43
?-domain structures 4-helix bundles
44
All-? proteins Globins
45
?-domain structures

• Anti-parallel ? structures
• Functionally most diverse
• Includes
• Up-and-down sheets or barrels
• Propeller-like structures
• Jelly roll barrels (from Greek key motifs)

46
Up-and-down sheets and barrel
47
Up-and-down sheets can form propeller-like
structures
48
Greek key motifs can form jelly roll barrels
49
a??-domain structures

• 3 main classes
• TIM barrel Core of twisted parallel strands
  close together
• Rossman fold open twisted sheet surrounded by
  helices on both sides
• Leucine-rich motif specific pattern of Leu
  residues, strands form a curved sheet with
  helices on outside

50
TIM barrel Rossman fold
51
Leucine rich motifs can form a???horseshoes
52
Protein structure databases visualization
software

• PDB Protein Data Bank
• http//www.rcsb.org/pdb/
• (RCSB) - several structure viewers
• MMDB Molecular Modeling Database
• http//www.ncbi.nlm.nih.gov/entrez/query.fcgi?db
  Structure
• (NCBI Entrez) - Cn3D viewer
• MSD Molecular Structure Database
• http//www.ebi.ac.uk/msd
• Especially good for interactions, binding sites
• Other visualization tools PyMol JMol

53
Protein structure classification

• SCOP Structural Classification of Proteins
• Levels reflect both evolutionary and structural
  relationships
• CATH Classification by Class, Architecture,
  Topology and Homology
• DALI/FSSP
• Fully automated structure alignments

For links discussion, comparisons of these,
see http//pdomains.rcsb.org/pdomains/index.php
54
Protein sequence databases

• UniProt (SwissProt, PIR, EBI)
• http//www.pir.uniprot.org
• NCBI Protein http//www.ncbi.nlm.nih.gov/entrez/
  query.fcgi?dbProtein

55
Protein sequence structure analysis

• Diamond STING Millennium - many useful structure
  analysis tools, including Protein Dossier
  http//trantor.bioc.columbia.edu/SMS/
• SwissProt (UniProt)
• knowledgebase
• http//us.expasy.org/sprot
• InterPRO
• sequence analysis tools
• http//www.ebi.ac.uk/interpro

56
Structural Genomics

• 20,000 "traditional" genes in human genome
• (not including miRNAs, etc.)
• 3,000 proteins in a typical cell
• gt 3 million sequences in UniProt
• 40,000 protein structures in the PDB
• Experimental determination of protein structure
  lags far behind sequence determination!
• Goal of Structural Genomics Determine structures
  of "all" protein folds in nature, using
  combination of experimental structure
  determination methods (X-ray crystallography,
  NMR, mass spectrometry) structure prediction

57
Structural Genomics Projects
TargetDB database of structural genomics
targets http//targetdb.pdb.org
Protein Structure Prediction?
58
Protein Folding

• "Major unsolved problem in molecular biology"
• In cells spontaneous
• assisted by enzymes
• assisted by chaperones
• In vitro many proteins fold spontaneously
• many do not!

59
Steps in Protein Folding

• 1- "Collapse"- driving force is burial of
  hydrophobic aas
• (fast - msecs)
• 2- Molten globule - helices sheets form, but
  "loose"
• (slow - secs)
• 3- "Final" native folded state - compaction, some
  2' structures
  rearranged
• Native state? - assumed to be lowest free energy
• - may be an ensemble of structures

60
Protein Dynamics

• Protein in native state is NOT static!
• Function of many proteins depends on
  conformational changes, sometimes large,
  sometimes small
• Recall
• Globular proteins are inherently "unstable"
• (most proteins have NOT evolved for maximum
  stability)
• Energy difference between native and denatured
  state is very small (5-15 kcal/mol)
• (this is equivalent to 1 or 2 H-bonds!)
• So Assumption of prediction methods that lowest
  free energy structure is "native" doesn't help a
  lot! There may be many "decoy structures" with
  very similar "energy" scores

61
Protein Structure Prediction

• Structure is largely determined by sequence
• BUT
• Similar sequences can assume different
  structures
• Dissimilar sequences can assume similar
  structures
• Many proteins are multi-functional
• 2 Major Protein Folding Problems
• 1- Determination of folding pathway
• 2- Prediction of tertiary structure from
  sequence
• Both still largely unsolved problems