Bioinformatics Seminar
PhD Alfonso Trezza
Doctorate in Molecular Biology and Biochemistry Department of Biotechnology, Chemistry and Pharmacy
University of Siena
E-mail: trezzaalfonso@gmail.com
TRIP
Structural Biology
About myself…
Education Nov 2015 - to date
PhD in Biochemistry and Molecular Biology, University of Siena Nov 2013 - Sep 2015
Master’s Degree in Cellular and Molecular Biology, University of Siena, 110 upon 110 summa cum laude.
Thesis title: “Innovative approach for the individuation of protein-protein interfaces active site potential involved in the onset of metastasis and rational small molecule disruptors drug design”
Sep 2009 - Sept 2013
Bachelor’s Degree in Biological Science, University of Salerno
Thesis title: “Linezolid and its analogues: Syntheses and mechanisms of action”
Publications
1. Alfonso Trezza, Vittoria Cicaloni, Piera Porciatti, Andrea Langella, Fabio Fusi, Simona Saponara, Ottavia Spiga. From in Silico to in Vitro: a trip to reveal Flavonoid Binding on the Rattus norvegicus ATP-sensitive inward rectifier potassium channel 8 (Kir6.1). PeerJ Article
2. PN Khanh, TT Huong, O Spiga, A Trezza, TD Cuong, VT Ha, NM Cuong. In silico screening of anthraquinones from Prismatomeris memecyloides as novel phosphodiesterase type-5 inhibitors (PDE-5Is). Revista Internacional de Andrología. 2017
3. Federico Galvagni, Federica Nardi, Ottavia Spiga, Alfonso Trezza, Giulia Tarticchio, Rosanna Pellicani, Eva Andreuzzi, Elena Caldi, Paolo Toti, Gian Marco Tosi, Annalisa Santucci, Renato V Iozzo, Maurizio Mongiat, Maurizio Orlandini. Dissecting the CD93-Multimerin 2 interaction involved in cell adhesion and migration of the activated endothelium. Matrix Biology 64, 112-127. 2017
4. Alfonso Trezza, Andrea Bernini, Andrea Langella, David B Ascher, Douglas EV Pires, Andrea Sodi, Ilaria Passerini, Elisabetta Pelo, Stanislao Rizzo, Neri Niccolai, Ottavia Spiga. A Computational Approach From Gene to Structure Analysis of the Human ABCA4 Transporter Involved in Genetic Retinal Diseases. Investigative ophthalmology & visual science 58 (12), 5320-5328. 2017
5. Fabio Fusi, Alfonso Trezza, Ottavia Spiga, Giampietro Sgaragli, Sergio Bova. Cav1. 2 channel current block by the PKA inhibitor H-89 in rat tail artery myocytes via a PKA-independent mechanism: Electrophysiological, functional, and molecular docking studies. Biochemical pharmacology 140, 53-63. 2017
6. F Fusi, O Spiga, A Trezza, G Sgaragli, S Saponara. The surge of flavonoids as novel, fine regulators of cardiovascular Cav channels. European journal of pharmacology 796, 158-174. 2017
7. A Trezza, A Bernini, O Spiga. Identification of Inhibitors Binding Site of Ebola L Polymerase Based on its Homology Model Virology & Antiviral Research 2016
8. PN Khanh, O Spiga, A Trezza, YH Kim, NM Cuong. Coumarins Isolated from Murraya paniculata in Vietnam and Their Inhibitory Effects against Enzyme Soluble Epoxide Hydrolase (sEH).. Planta Medica International Open 3 (03), e68-e71. 2016
9. F Fusi, M Durante, O Spiga, A Trezza, M Frosini, E Floriddia, E Teodori, S Dei, S Saponara. In vitro and in silico analysis of the vascular effects of asymmetrical N, N-bis (alkanol) amine aryl esters, novel multidrug resistance-reverting agents. Naunyn-Schmiedeberg's archives of pharmacology 389 (9), 1033-1043
10. A Trezza, A Bernini, O Spiga. Identification of “on-off residues” in rat Cav1. 2 α1C subunit channel using in silico analysis and docking simulation.. PeerJ Preprints
11. Co-Author of a chapter “A Focus on Ebolavirus Polymerase: Structure, Functions and Antiviral Therapies” of the book: VIRAL POLYMERASES, STRUCTURES, FUNCTIONS AND ROLES AND ANTIVIRAL DRUG TARGET (the “Work”) edited by Satya Prakash Gupta (the “Editor”). This Agreement between you (the “Contributor”) and Elsevier Inc. (the
“Publisher”)
Where and How do proteins get to be born?
Where? How?
How it’s made a Protein?
A Protein is made up by aminoacids (small organic molecules)
There are only 20 aminoacids
The variety of proteins is given from the
combination of aminoacids
A protein of n residues has 20n possible sequence
What's Protein shaped like?
The proteins have 4 structures:
1. Primary structure (aminoacids sequence) 2. Secondary
structure ( α-helix, β-sheet and loop composition) 3. Tertiary structure
(α-helix, β-sheet and loop three- dimensional structure) 4. Quaternary
structure ( tertiary structure
aggragation)
How many kinds of proteins are there?
There are lots of protein kinds, but we can split it in 2 macro families:
Soluble Proteins
(Free inside the cytoplasm) Membrane Proteins
(Inside the cell membrane)
What’s shapes are the proteins?
The proteins can have a several structures, according to the composition of aminoacids
On the basis of the folding the proteins can have different funticons
If the protein is not well folded, it will be destroyed
Some protein structures are very funny!!!
How big are the proteins?
http://learn.genetics.utah.edu/content/cells/scale/
From the DNA to the Protein
From DNA to protein - 3D.mp4
Why to study the proteins?
Cancer
Cardiovascular diseases
Metabolic diseases
Cerebral diseases
To know the protein structures is a crucial step in order to identify potential drugs against these «bad proteins»
…But it is fundamental to see the protein…
Can I see the proteins? Yes
There are 3 methods principally:
1. N.M.R (Nuclear magnetic resonance) 2. Ray-x
3. Cryo-em
N.M.R Ray-x
Cryo-em
But…
where are the proteins???
Protein Data Bank (PDB)
CXCL12
Can I see the proteins? Yes
There are lots a tool able to show the proteins, one of the best is PyMOL
It’s Free and OpenSource
Version Avaible for Windows, Mac and Linux
Want you to see how PyMOL works?
The key is the: @ !£$%&/(=?^#§>
1. Can we do a protein structure ex novo?
2. Can we look for a potential drug of a protein?
3. Can we see how the potential drug operates against the protein?
Yes, we can!!!
The way is only one, The key is the:
BIOINFORMATICS
Can we do a protein structure ex novo? Yes, the Homology Modeling is the way
The homology modeling is a bioinformatics approach which let us model a protein structure ex-novo…
How can we do a protein structures from their primary structures?
(The alignment of sequences is the way)
Similarity = 60%
? ?
How can we identify a potential drug against a protein?
(The docking simulation is the way)
The docking simulation is a protein-molecule interaction simulation
The simulation can be carried out through different tool (free or payment)
The time of
simulation is different according of
tools( from few seconds to days)
The final result can be different
(chemical-physical propriety, molecular size, ect.)
Name_molecule Docking Score
1 Score 1
2 Score 2
3 Score 3
4 Score 4
The docking simulation
Disco Docking - Computational Drug Design.mp4
We have a problem!!! The proteins are always in motion… Right now
The «breath» of the protein involve different considerations:
1. The potential binding site ( where is the drug ) could modify its conformation, decreasing the interaction with the drug
2. A region of the protein (originally covered up) could become a potential binding site (transient pocket)
3. The binding of the drug could modify the protein conformation, allowing a second interaction of an other drug
4. The breath of the protein could get out the drug from the binding site
So…
Can we simulate the moviment of a Protein? Of course
(The Molecular Dynamics Simulation is the way)
“In the real world, this could eventually mean that most chemical experiments are conducted inside the silicon of chips instead of the glassware of laboratories. Turn off that Bunsen burner; it will not be wanted in ten years.”
- The Economist, reporting on the work of the 1998 Chemistry Nobel Prize Awardees
* Many Physically-Based Simulations model easily observable real world
phenomena.
* Molecular Dynamics
Simulations model things too small for us to
observe directly.
• Modeling the motion of a complex molecule by solving the wave functions of the various subatomic particles would be accurate…
• But it would also be very hard to program and take more computing power than anyone has!
Why Not Quantum Mechanics?
) , , ( )
, , ( )
, , 2 (
2 2
z y x E
z y x z
y x
m U
Classical Mechanics
Instead of using Quantum mechanics, we can use classical Newtonian mechanics to model our system.
This is a simplification of what is actually going on, and is therefore less accurate.
To alleviate this problem, we use numbers derived from QM for the constants in our classical equations
Molecular Modeling
For each atom in every molecule, we need:
Position (r)
Momentum (m + v)
Charge (q)
Bond information (which atoms, bond angles, etc.)From Potential to Movement
To run the simulation, we need the force on each
particle.
We use the gradient of the potential energy function.
Now we can find the acceleration.
i i
i m a
F
V F i i
2 2
dt r m d
dr
dV
ii i
What is the Potential?
A single atom will be affected by the potential energy functions of every atom in the system:
Bonded Neighbors
Non-Bonded Atoms (either other atoms in the same molecule, or atoms from differentmolecules)
bonded non
bonded E E
R
V ( )
Non-Bonded Atoms
There are two potential functions we need to be concerned about between non-bonded atoms:
van der Waals Potential
Electrostatic Potentialtic electrosta Waals
der van
bonded
non
E E
E
The Electrostatic Potential
Opposite Charges Attract
Like Charges Repel
The force of the attraction is inversely proportional to the square of the distance
pairs
nonbonded ik k tic i
electrosta
Dr
q
E q
This is the Molecular Dynamics Simulation
Simulation of millisecond protein folding NTL9 (from Folding@home).mp4
The last frontier of Molecular Dynamics:
SuperVised Molecular Dynamics (SuMD)
The major constraint of docking simulation is the rigidity of the system ( the protein and the drug are still).
So…
Can we simulate if a drug is able to bind against a protein?
Can we simulate as bind a drug inside the protein?
Can we simulate how many time a drug binds inside the protein?
Can we see all this?
Yes, We Can…The SuMD is the way…
ci5b00702_si_002.mpg ci5b00702_si_003.mpg ci5b00702_si_008.mpg
Inner Life Of A Cell - Full Version.mp4
