Structural Biochemistry
Code
7637
Academic unit
Faculdade de Ciências e Tecnologia
Department
Departamento de Química
Credits
6.0
Teacher in charge
Maria dos Anjos Lopes de Macedo, Maria João Lobo de Reis Madeira Crispim Romão
Weekly hours
8
Teaching language
Português
Subject matter
Part I
- Basic Concepts in NMR (review); 1D and 2D spectra (homo-and hetero-nuclear): applications to proteins
- Identification and assignment of signals in protein 2D spectra; secondary structural elements determination based on chemical shifts
- Backbone assignment of proteins using triple resonance experiments - methodology
- Determination of protein structures in solution; NMR constraints and methodology for structure calculations
- Relaxation and protein dynamics
- NOE effect and Saturation Transfer Difference experiments: application to protein - ligand interactions
- NH exchange and protein structural information.
Part II
- Introduction to protein crystallography; review
- Crystallization of proteins; Lattice, Symmetry and Space Groups
- X-ray sources and detectors; diffraction; reciprocal space
- Diffraction and Fourier transform; The Phase Problem and how to solve it.
- Model building and Refinement; Methods of Validation
- Molecular Electron Microscopy
Practical Classes
NMR solution structure
- P1 : The NMR spectrometer ; 2D heteronuclear experiments.
- Homework / Q1: Determination of the secondary structure of a protein based on the amino acid sequence ( eg CBM11 ); online tools .
- P2 : Resonance assignment using 3D spectra. Introduction to BMRB - Biological Magnetic Resonance Data Bank.
- Homework / Q2 : Secondary structural elements determination based on backbone chemical shifts.
- P3 : Solution structures determination using Cyana software - input data preparation, output data (violations detection & iterative process) and validation results (online tools). NMR vs X-ray structures.
- Homework / Q3 : NMR structures validation (using online tools)
- P4 : Protein ligand interactions; titrations followed by HSQC and data obtained by STD (eg CBM11 )
- Homework / Q4 : delivery of a written analysis of a scientific article - case study on protein - ligand interactions
- S : Case study Oral presentation and discussion
X-Ray Crystallography
- P1: databases, PDB revisited
- P2: Preparation and optimization of protein crystals (Q1)
- P3: The X-ray diffractometer and diffraction experiment. Processing and analysis of data.
- P4: Solving a structure by MR method (Q2)
- P5: Analysis of electron density and model building; Model quality and validation criteria. (Q3)
- S: Seminars – Case study : Oral presentation and discussion