Guide to PISA

Getting started

PISA stands for Protein Interfaces Surfaces and Assemblies. It is available as a webservice at the EBI, via:

• http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html

PISA is a tool for exploring macromolecular interfaces and surfaces, and for examining probable quaternery structure (also referred to as "assemblies"). It can do this using either precalculated data from the PDB archive, or coordinate data uploaded by the user.

The analyses are:

• structural/chemical properties of macromolecular surfaces
• structural/chemical properties of interfaces
• probable assemblies), their structural/chemical properties and probable dissociation pattern

Other functions include: searching the PDB archive for interfaces formed by structural homologues; assessing the biological role of interfaces; and the ability to download and visualise the structures, interfaces and assemblies using a variety of software.

Use 3GCB as an example

The initial PISA start page has the 3GCB structure preselected so I will used this as an example to explore the use of PISA.

3GCB is GAL6 (YEAST BLEOMYCIN HYDROLASE) MUTANT C73A/DELTAK454. Once the accession code has been entered PISA will give you some basic starting information on the page, namely:

• The number of chains and ligands in the entry (e.g. 1 aminoacid chain and 7 ligands in ASU), and
• The most probable assembly (e.g. 6-mer).

The chains and ligands form the basic components of the "structures" that are used in the analyses later on. So for example in the 3GCB example, the structures are the main chain and the seven ligands: GOL residues 1001 and 1002, and SO4 residues 1101 to 1105.

You are then presented with a choice of three different aspects of the protein to explore:

• interfaces
• structures
• assemblies

Structures

This is probably a good place to start. The first section Interfacing Structures lists in a table the different "structures" found within the entry, between which interfaces can form.

Each structure is given a row number (in the column marked "NN") which links to the details of that structure (the rows in the table are grouped according to some scheme which isn't clear to me).

A structure is described by:

• range of residues (or {-} to indicate the main chain)
• class, which is one of Protein (amino acid chains), DNA/RNA (nucleic acid chains) or Ligand (small molecules which are not solvent).

Other details of the structure include the numbers of atoms (N_at) and residues (N_res), and information about the surface of the structure:

• Number of surface atoms (^SN_at) and residues (^SN_res)
• The solvent accessible surface area

If applicable the value of ΔG (the solvation free energy of folding for the structure) is also shown.

At the bottom there is a table of structure similarity. This gives an indication of which structures within the entry are similar to any other structures - a cyan block in the matrix indicates that the two structures are similar (black indicates that they are the same!), and clicking on the dot allows you to see the two structures superposed.

Interfaces

Interfaces between pairs of "structures" (i.e. chains and ligands) within the entry.

A results table shows details of each interface found, with one interface per row of the table:

• Structure 1 is defined as a range of residues or as as a ligand name. {-} indicates the whole chain; other structures are the ligands and hetatm groups indicated as e.g. [GOL]-:1001.
• Structure 2 is the second component which interfaces with the first. It is defined in a similar way to structure 1, however an additional symmetry operation is supplied (expressed either as a string or as a code).

For each structure the number of interfacing atoms (ⁱN_at) and residues (ⁱN_res) are indicated.

For each interface a number of quantities are listed:

• Interface area (in A²)
• ΔⁱG expressed in kcal/M, and an associated "P-value". ΔⁱG is the solvation free energy gain on formation of the interface, with negative values indicating a hydrophobic interface.
  The P-value is a probability measure of the specificity of the interface:
  • P=0.5 indicates that the ΔⁱG value is typical for this type of interface.
  • P>0.5 means that the interface is less hydrophobic than expected, suggesting that it is an artefact of crystal packing.
  • P<0.5 indicates interfaces with an unexpectedly large hydrophobicity, and thus a high specificity.
• The number of potential hydrogren bonds (N_HB) and salt bridges (N_SB) across the interface
• The "complexation significance score" CSS, which scores how significant the interface is for assembly formation.

A link (in the column labelled "x") launches the selected viewer to visualise the interface. Clicking on the interface number (in the column marked "NN") links to more detail about the interface in question.

Assemblies

This page gives information about the possible quaternery structures ("assemblies") that PISA predicts. The table summarises the predicted assemblies, and clicking on the id number goes to the details for that assembly.

The summary for each assembly consists of the following information:

• mmSize is the number of macromolecular monomeric units in each assembly (consisting of protein and/or RNA/DNA, but not ligands), and the number of assemblies in the unit cell. For example: "2x6" indicates that the assembly contains 6 macromolecular monomeric units, and, for the given PQS set, a unit cell contains 2 assemblies of this type.
• Formula and composition give respectively the chemical make up of the assembly. Note that the formula doesn't use the chain ids from the entry, whereas the composition does.
• Details of solvent accessible and buried surface areas, and the solvation free energy on formation (ΔG^int) and disassociation (ΔG^diss) of the assembly.

Notation for structures

Structures are indicated using the following notation.

Full chains without a chain ID are denoted {-}. Ligands are denoted:

[R]C:N

where R is the residue or ligand name, C is the chain, and N is the residue number. For example, [HEM]-:605 means protoporhyrin IX containing Fe (HEM) in chain without a chain ID, residue number 605.