EMBL-EBI

MSD database is structured around the fact that Proteins are “sticky”

EMBL-EBI

A short biography of 1 protein whose very existence depends on being as sticky as possible

EMBL-EBI

EMBL K02078

ttaacgcgta aattcaaaaa tctcaaattc cgacccaatc aacacacccg ataccccatg ccaataaaaa agtaacgaaa atcggcacta aaactgacaa ttttcgacac tgccgccccc ctacttccgc aaaccacacc cacctaaaag aaaatacaaa ataaaaacaa ttatatagag ataaacgcat aaaatttcac ctcaaaacat aaaatcggca cgaatcttgc tttataatac gcagttgtcg caacaaaaaa ccgatggtta aatacattgc atgatgccga tggcaagccc tgaggctttc ccctttcaat taggagtaat tttatgaata cccttcaaaa aggctttacc cttatcgagc tgatgattgt gatcgctatc gtcggcattt tggcggcagt cgcccttccc gcctaccaag actacaccgc ccgcgcgcaa gtttccgaag ccatcctttt ggccgaaggt caaaaatcag ccgtcaccga gtattacctg aatcacggca aatggccgga aaacaacact tctgccggcg tggcatcccc cccctccgac atcaaaggca aatatgttaa agaggttgaa gttaaaaacg gcgtcgttac cgccacaatg ctttcaagcg gcgtaaacaa tgaaatcaaa ggcaaaaaac tctccctgtg ggccaggcgt gaaaacggtt cggtaaaatg gttctgcgga cagccggtta cgcgcaccga cgacgacacc gttgccgacg ccaaagacgg caaagaaatc gacaccaagc acctgccgtc aacctgccgc gataaggcat ctgatgccaa atgaggcaaa ttaggcctta aattttaaat aaatcaagcg gtaagtgatt ttccacccgc ccggatcaac ccgggcggct tgtcttttaa gggtttgcaa ggcgggcggg gtcgtccgtt ccggtggaaa taatatatcg at

EMBL-EBI

MNTLQKGFTL IELMIVIAIV GILAAVALPA YQDYTARAQV SEAILLAEGQ KSAVTEYYLN HGKWPENNTS AGVASPPSDI KGKYVKEVEV KNGVVTATML SSGVNNEIKG KKLSLWARRE NGSVKWFCGQ PVTRTDDDTV ADAKDGKEID TKHLPSTCRD NFDAK

UniProt P02974

EMBL-EBI

PDB 1AY2

MSD DATABASE

pentamer

MSD DATABASE

negatively stained TEM images

Neisseria gonorrhoeae expressing pili and interacting with epithelial cells. The pili are polar flexible filaments of about 5.4 nm diameter and 2500 nm average length.

Type IV Pilin Structure and Assembly: X-Ray and EM Analyses of Vibrio cholerae Toxin-Coregulated Pilus and Pseudomonas aeruginosa PAK Pilin

L. Craig, et al Molecular Cell, 11, 1139–1150, 2003

EMBL-EBI

Type IV pili are not merely passive sticky fibres but dynamic machines that participate in a surprising number of functions including:

Bacterial aggregation Adhesion to host cells Twitching motilityPilus retraction DNA transformation In another bacterial species, motility. Phage receptor in V. cholerae.

EMBL-EBI

EMBL

UniProt

PDB

Assembly (MSD)

Microscopy

still not the full story - GENOME

EMBL-EBI

Pilus gene organisation

Many copies of pilin gene throughout chromosome

Two are functional, pilE1 and pilE2 All other copies are silent Antigenic variation occurs due to

recombination (within mini-cassettes)

EMBL-EBI

Antigenic variation in N. gonorrhoeae

A single cell can give rise to daughter cells expressing structurally and antigenically different pili

Gonococcus has the genetic capacity to make as many as a million different pilin variants

All able to bind to same host tissues and to cause the same disease symptoms

EMBL-EBI

PDB Entries and X-Ray results

1. Crystal Structure

2. Molecular Structure (covalent)

3. Oligomeric Assembly

What has all this got to do with MSD?

EMBL-EBI

Chains Residues AtomsExp. Result Assembly

ALT

ASSEMBLY

ASSEMBLY DATA

ATOM

ATOM DATA

CHAIN

COMPONENT

DEPOSITION

MODEL

atd_component_fkatd_component_fk

assembly_deposition_fk

assembly_deposition_fk

assembly_a_data_fk

assembly_a_data_fk

assembly_data_model_fk

assembly_data_model_fk

atd_alt_fk

atd_alt_fk

atd_atom_fk

atd_atom_fk

atd_chain_fk

atd_chain_fk

chain_assembly_fk

chain_assembly_fk

atd_model_fk

atd_model_fk

MSD Relational Database

EMBL-EBI

KEY to MSD DataBase

EMBL-EBI

Biological Context

PDB MSD

Oxalate oxidase 1FI2 hexameric

EMBL-EBI

PDB Xray coordinates

PDB entry the deposited coordinates usually consist of the contents of the asymmetric unit:

The contents of the ASU define a single copy of the macromolecule

The contents of the ASU consist of more than one copy of the macromolecule

The contents of the ASU require crystallographic symmetry operations to be applied to generate the complete macromolecule(s)

A combination of the above, including multiple copies and required symmetry transformations

EMBL-EBI

benzene

C6H6

Covalent bonded

EMBL-EBI

Benzene crystallised in

Space Group P6/m

6-fold rotation axis

Mirror plane

EMBL-EBI

Benzene P6/m in the PDB

ATOM C1 x1 y1 z1 occupancy 0.5

ATOM H1 x2 y2 z2 occupancy 0.5

Entire atomic contents:

EMBL-EBI

The stronger of the two is the hydrogen bond.

The weaker is the van der Waal's forces.

Both interactions depend on the same fundamental cause, the charge on electrons, and how that results in attraction and repulsion at an atomic level.

HELD TOGETHER BY WEAK FORCES

EMBL-EBI

Quaternary Structure

Quaternary Structure is defined as that level of form in which units of tertiary structure aggregate to form homo- or hetero-multimers.

Consideration of the presence of a quaternary state is important in the understanding of a protein's biological function.

Crystal Structure

Crystal Structure

Oligomeric Assembly

EMBL-EBIProteins don’t do this –

pack by translationals

EMBL-EBI

There are three main types of symmetry:

symmetry with respect to a plane (mirrors) symmetry with respect to a line (rotations) symmetry with respect to a point (inversions)

Symmetry

EMBL-EBI

symmetry with respect to a line (rotations)

symmetry with respect to a plane (mirrors) symmetry with respect to a point (inversions)

Symmetry

EMBL-EBI

1, 2, 3, 4, 6 -fold rotational symmetry

These are the only rotational symmetries that can exist in crystals; all others are disallowed. These five rotational axes are called the five Proper Axes

Symmetries showing 5-, 7-, 8-, 9-, 10-, 11-, & 13- fold rotations are known for biological molecules – these are observed in the Asymmetric Unit.

Rotational symmetry

1g8h

Applying 1st 3-fold Rotation

A

A’

Residues of Chain A in interface

A

A’ Residues of Chain A’ in interface

Applying 2nd 3-fold Rotation

A

A’

A”

Also has a 2-fold rotation

Final Assembly is a Hexamer from 23 symmetry

EMBL-EBI

If you add translations to rotation axes, you form what are call screw axes. For an nm screw axis, the rotational component is 360/n degrees, and the translations is m/n of the unit translation along the axis.

In Biological Crystallography --> Polymers

Helices are improper Screw axes – e.g. DNA

Screw Axes

EMBL-EBI

Also has a 2-fold rotation – infinite cylinder in crystal

Screw Axis

EMBL-EBI

Screw Axes example

tubulins

EMBL-EBI

SYMMETRY Rules –BUT What about -

What happened to symmetry?

2:1 hetero-complex

The Ribosome – the champion Heterocomplex

proteins tossed around the RNA

protein aggregates complicate the lives of people who study proteins in vitro

Protein Aggregation and Amyloid Diseases- Converting the protein from a soluble to a fibrillar structure