A cell is an organization of millions of molecules

A cell is an organization of millions of moleculesA cell is an organization of millions of molecules

Proper communication between these molecules Proper communication between these molecules is essential to the normal functioning of the cell is essential to the normal functioning of the cell

To understand communication between To understand communication between molecules:molecules: *determine the arrangement of the atoms*

Organ Organ Tissue Tissue Cell Cell Molecule Molecule Atoms Atoms

Structural BiologyStructural BiologyMedicine and Biology at the Atomic ScaleMedicine and Biology at the Atomic Scale

High Resolution Structural BiologyHigh Resolution Structural Biology

Using atomic structure to understand how Using atomic structure to understand how biomolecules communicatebiomolecules communicate

Anti-tumor ActivityDuocarmycin SA

The Reward: UnderstandingThe Reward: UnderstandingControlControl

Shape

Atomic interactions

Hierarchy of Structural BiologyHierarchy of Structural Biology

MoleculeStructural Genomics

PathwayStructural Proteomics

ActivitySystems Biology

RPARPANER

BER

RR

High Resolution Structural BiologyHigh Resolution Structural BiologyThree Complementary TechniquesThree Complementary Techniques

NMR Spectroscopy X-ray Crystallography

Computation

Determine experimentally or modelDetermine experimentally or model3D structures of biomolecules3D structures of biomolecules

What Tools Do We Use?What Tools Do We Use?

NMRNMR ComputationComputation

RPA-B

RPA-AB

RPA-A

Fluo

resc

ence

Inte

nsity

Ratio of T-ag/RPA

Biophysical AnalysisBiophysical AnalysisExpression/MutationsExpression/Mutations CrystallographyCrystallography

Experimental DeterminationExperimental Determination of 3D Structures of 3D Structures

X-ray

X-raysDiffraction

Pattern

Direct detection ofatom positions

Crystals

NMR

RFRF

Signals

H0

Indirect detection ofH-H distances

In solution

Representations of StructureRepresentations of Structure Conformational EnsembleConformational Ensemble

PRECISIONRMSD of the ensemble

Neither crystal nor solution structures are properly represented by a single conformation Intrinsic motions Imperfect data

Computational Determination Computational Determination of 3D Structuresof 3D Structures

• Specify the forces that act on each atom

• Simulate these forces on a molecule of known atomic composition

• Start with experimental data or approximate experimental structure or just sequence

• Many algorithms in use, all treatments for bio-macromolecules are empirical

Unique Contributions of Each Unique Contributions of Each Structural Biology TechniqueStructural Biology Technique

• X-ray- highest resolution, automation coming

• NMR- enables wide variation of solution conditions; direct characterization of motions and of weakly interacting systems

• Computation- fundamental understanding of structure, dynamics and interactions; models without experiment; fast

Challenges For Understanding Challenges For Understanding The Meaning of StructureThe Meaning of Structure

• Structures determined by NMR, computation, and X-ray crystallography are static snapshots of highly dynamic molecular systems

• Biological process (recognition, interaction, chemistry) require molecular motions (from femto-seconds to minutes)

• New methods are needed to comprehend and facilitate thinking about the dynamic structure of molecules: visualization

Visualization of StructuresVisualization of Structures

Intestinal Ca2+-binding protein!

Need to incorporate 3D and motion

Calcium-Dependent Protein KinaseCalcium-Dependent Protein KinaseA Target for Anti-Malarial Drug DesignA Target for Anti-Malarial Drug Design

Ca2+

Target(kinase)

Calcium Signal TransductionCalcium Signal Transduction

EF-Hand Ca2+ Sensors

Regulation of Protein Kinase Regulation of Protein Kinase Activity by CaActivity by Ca2+2+-Calmodulin -Calmodulin

CaCa2+2+ signal signal

Kinase ActivationKinase Activation

Protozoa Have A Unique Kinase Protozoa Have A Unique Kinase The CaThe Ca2+2+-Dependent Protein Kinase (CDPK)-Dependent Protein Kinase (CDPK)

Kinase

CDPK has its own regulatory apparatus!!How does it work?

Standard auto-inhibitory segment

Calmodulin-like domain (CaM-LD)

Structure of CDPK RevealsStructure of CDPK RevealsIntra-molecular Kinase ActivationIntra-molecular Kinase Activation

Anti-Malarial Drug DesignAnti-Malarial Drug DesignMust Inhibit CaM-LD But Not CalmodulinMust Inhibit CaM-LD But Not Calmodulin

Calmodulin CaM-LD

Center for Structural BiologyCenter for Structural BiologyA Trans-Institutional InitiativeA Trans-Institutional Initiative

•Increase molecular/physical thinking in medical and biological research•Foster multi-disciplinary approaches•Focal point bridging medicine and biology to physics and chemistry

Structural Biology ResourceStructural Biology ResourceBut Not A Core!!But Not A Core!!

• Education/project origination• Enabling expertise: 3 Ph.D. Staff

Scientists• Enabling hardware: technologies to

visualize and determine structures• Open-access focal point in BIOSCI/MRBIII