A 7th EC Class for Translocases

Six enzyme classes have been recognized since the first Enzyme classification and nomenclature list was first approved by the International Union of Biochemistry in 1961. These were based on the type of reaction catalyzed:

Oxidoreductases (EC 1)
Transferases (EC 2)
Hydrolases (EC 3)
Lyases (EC 4)
Isomerases (EC 5)
Ligases (EC 6)

However, it has become apparent that none of these could describe the important group of enzymes that catalyse the movement of ions or molecules across membranes or their separation within membranes. Several of these involve the hydrolysis of ATP and had been previously classified as ATPases (EC 3.6.3.-), although the hydrolytic reaction is not their primary function.

These enzymes have now been classified under a new EC class of Translocases (EC 7). The reactions catalyzed are designated as transfers from ‘side 1’ to ‘side 2’ because the designations ‘in’ and ‘out’ (or ‘cis’ and ‘trans’), which had been used previously, lack clarity and can be ambiguous. The comments associated with each entry then describe the specific translocations catalysed.

The subclasses designate the types of ion or molecule translocated:

• EC 7.1 contains enzymes catalysing the translocation of hydrons (hydron being the general name for H⁺ in its natural abundance),
• EC 7.2 contains those catalysing the translocation of inorganic cations and their chelates,
• EC 7.3 contains those catalysing the translocation of inorganic anions,
• EC 7.4 contains those catalysing the translocation of amino acids and peptides,
• EC 7.5 contains those catalysing the translocation of carbohydrates and their derivatives
• EC 7.6 contains those catalysing the translocation of other compounds.

The sub-subclasses concern the reaction that provides the driving force for the translocation, where these are relevant:

• EC 7.x.1 translocations linked to oxidoreductase reactions
• EC 7.x.2 translocations linked to the hydrolysis of a nucleoside triphosphate
• EC 7.x.3 translocations linked to the hydrolysis of a diphosphate
• EC 7.x.4 translocations linked to a decarboxylation reaction

Exchange transporters that are not dependent on enzyme-catalyzed reactions, such as the exchange of ions across membranes, are not included. Pores that change conformation between open and closed states in response to phosphorylation or some other catalyzed reaction are classified under EC 5.6 (Macromolecular conformational isomerases).

(prepared by Keith Tipton, originally published at the ExplorEnz enzyme database)

Pan-Genome PGDBs Unify Genomic and Metabolic Data across Related Strains

Pan-Genome PGDBs are a relatively new feature of BioCyc. These BioCyc databases combine in one place information about multiple sequenced genomes for a given species. For example, the Helicobacter pylori Pan-Genome database covers 158 sequenced strains.

The Pan-Genome PGDBs contain one gene object for each orthologous group of genes in the organism. They also contain the union of all metabolic pathways across all the strains. Thus, a Pan-Genome PGDB allows you to quickly assess the full set of gene functions and metabolic pathways across the the known strains.  For example, the gene page shows all orthologs across all strains in the Pan-Genome.  The page for the ftsX gene illustrates this for a gene with relatively few orthologs.

Other gene pages list hundreds of orthologs and synonyms.

You can visit a page for the orthologous genes by following the links in the 'Relationship Links' area near the bottom of the page.

Pan-Genomes provide a way to visualize genes in the genome browser as well.   It's easier to understand this visualization if you know that Pan-Genomes are constructed starting with the PGDB of a base strain and adding other members of the collection of strains one-by-one.  For example, the H. pylori Pan-Genome was constructed by starting with strain 26695.  The visualization is based on dividing groups of orthologous genes into two sets: ortholog groups that include genes that occur in the base strain, and ortholog groups that only include genes in other strains.  

Ortholog groups that include genes from the base strain are collected on a 'chromosome' that preserves the location and direction of the genes from the base strain.  The remaining ortholog groups are mapped in arbitrary order onto an 'artificial replicon'.  The names in the artificial replicon display are based on the (arbitrary) order that PGDBs were added to the Pan-Genome.

You can search for genes either by name (e.g., 'abc') or an identifier combining a strain name and id joined with an underscore (e.g., HPHPP11_0013) by entering the name in the search box at the top right.  Either quick search or gene search will take you to the page containing the gene and its orthologs in the Pan-Genome PGDB. 

The cellular overview diagram provides a way to visualize reactions associated with genes shared by all members of the Pan-Genome. You can also visualize those reactions that are unique to a single organism within the Pan-Genome.  In the screen shot below, the reactions shared by all organisms in the H. pylori Pan-Genome are shown in red and those unique to a single organism are purple.

To create a diagram like this on BioCyc web site, select a Pan-Genome PGDB, and bring up the cellular overview. In the operations menu, choose Highlight Genes -> By Pan-Genome Core Genes.  The core genes are the set of genes shared by all organism databases in the Pan-Geneome.  Chose Highlight Genes -> By Pan-Genome Unique Genes, to highlight reactions associate with genes that are unique to a single organism database.

In the desktop, show the Cellular Overview, then from Overviews -> Highlight -> Highlight the core genome followed by Overviews -> Highlight the unique genes will achieve the same highlighting.


More details on how Pan-Genome PGDBs are created and how to use them are provided here.

You can select a Pan-Genome PGDB by entering the phrase “pan-genome” in the change organism database dialogue. Here's our current list of Pan-Genome PGDBs and the number of strains that each one contains. Over time we will be adding Pan-Genome PGDBs for additional species, and regenerating existing Pan-Genome PGDBs to include additional strains.

Clostridioides difficile 10 strains

Escherichia coli 374 strains

Helicobacter pylori 158 strains

Listeria monocytogenes 35 strains

Mycobacterium tuberculosis 24 strains

Pseudomonas aeruginosa 24 strains

Salmonella enterica 113 strains

Shigella flexneri 9 strains

Vibrio cholerae 81 strains

Generating a SmartTable of Orthologous Genes Across Multiple BioCyc Genomes

This post shows how, given a list of gene names and/or identifiers from one organism, to retrieve the orthologous genes in a second organism.    For example, this procedure could be used to find the orthologs in EcoCyc of a set of genes of uncertain function in another organism, potentially providing insights about their functions. 

The BioCyc project generates a database of orthologous genes between many of the organism Pathway Genome Databases (PGDBs) we maintain.  These orthologs are stored in a central MySQL database that is separate from the individual BioCyc PGDBs.  This post describes a way to retrieve orthologs for a list of genes using a SmartTable.  We don’t recommend generating a ortholog list for a whole genome because of performance problems that, as of June 2018, we are working to resolve.   

Since we will be using a PGDB in addition to EcoCyc, you will need a BioCyc subscription (not just a free account) to follow through this demonstration.  Here’s a screenshot of the final SmartTable, which you can access directly here. We will begin with a file of 88 genes from E. coli strain B str. REL606 and determine their orthologs in EcoCyc.  The columns in the resulting SmartTable are as follows:  the list of gene names from the input file, their ‘ECB’ accession IDs genes, the names of orthologous genes from EcoCyc, and  ‘b-number’ accession IDs from EcoCyc.  We also show how to add a column containing gene product names.

Here is the step-by-step procedure.

1.     Go to the bottom of this post and cut and paste the list of gene identifiers into a text editor such as textedit or atom.  Save the file as 88EcoliGenes.txt.

2.     Go to BioCyc.org and login and use change organism database to change your organism to Escherichia coli B str. REL606. 

3.     Open the ‘Smart Tables’ menu and choose the ‘My Smart Tables’ command.  It actually doesn’t matter which of the commands in the menu you choose.

4.     You’ll find the operations menu in the upper right corner of the SmartTables page.  Under the ‘New’ command, you’ll find ‘Smart Table from Uploaded File’.  Choose that command. 

5.     In the resulting pop-up window, click the ‘Choose File’ button and select the file you saved out in step 1.  Once you have located and selected the file, review the options below the ‘Choose File’ button in the  upload window.  Since this is a file of gene identifiers, you should keep the ‘Try to make objects of type’ box checked, along with the radio button next to ‘Gene’.  Also leave the other two check boxes checked.  Click the ‘Upload’ button and a new SmartTable with a single column will appear.  If you see a warning message, ignore it and continue.

6.     Add accession numbers for the genes, by locating the ‘ADD PROPERTY COLUMN’ dropdown menu, then choose ‘Accession-1’ from the list.  You’ll see a list of ECB identifiers numbers’ for each of the genes on the first page. 

7.     Now that the table has some basic identifiers for the E. coli B str. REL606 genes, you can proceed to adding orthologs from EcoCyc.  Find the ‘ADD TRANSFORM COLUMN’, and choose the second transform ‘Compare – map to other species PGDB’.  That will bring up a pop-up list of species, which will be rather long.  Scroll down through the list until you find ‘Escherichia coli K-12 substr. MG1655’.  Once you have selected the right organism, click the ‘Go’ button.

8.     Now a column containing ortholog gene names appears.  Each entry is linked to the gene page in its organism’s PGDB.  Try it by clicking on thrL, and notice the organism in the upper left of the gene page.  Clicking on the browser back button will take you back to the table and restore the current PGDB to EcoCyc.

9.     To add a column containing accession numbers of each orthologous gene, select the ortholog column by clicking anywhere in the column header except where the title is displayed.   When the column is selected the header turns darker (and the ‘Gene Name’ column lightens as it is no longer the selected column).  Then, as in step 6,  choose ‘Accession-1’ from the dropdown below ‘Add Property Column.’  A different list of accession numbers appears.  Notice that both lists of accession numbers are simply strings that don’t link to any genes.  A column of product names for the orthologs can be added in a similar fashion.

10.  Finally, suppose you were interested only in those B strain genes having orthologs in the K-12 strain. To remove genes lacking orthologs, select the third column (‘Map to Escherichia…’).  Now look over to the operations menu on the right.  Select the ‘Filter’ command. A  pop-up window titled ‘Column text-value filter’ will appear. 

11.  This dialog has a series of options chained together in a ‘sentence.’  In the second drop-down, change ‘a copy of …’ to ‘this SmartTable’.  In the third option, choose ‘contain an object’.  Click the ‘Go’ button.  You should be left with a SmartTable containing 84 rows, the B / K-12 strain ortholog pairs, as shown in the screen capture at the top of this post.

12.  Finally, you can save the results to a file by selecting the Export command from the Operations menu.  Choose ‘to Spreadsheet file’ and since all of the genes have names in column one, you can save as common names rather than frame IDs, which are more readable in a spreadsheet.

As always, I welcome your questions and comments below.

Here is the example data.  Cut the area between the horizontal lines and paste into a text editor, such as textedit or atom, and save the file as 88Ecoligenes.txt.

---

thrL

thrA

thrB

thrC

yaaX

yaaA

yaaJ

talB

mog

satP

yaaI

dnaK

dnaJ

insL-1

mokC

hokC

nhaA

nhaR

ECB_00020

ECB_00021

ECB_00022

insA-1

insB-1

ECB_00025

ECB_00026

rpsT

yaaY

ribF

ileS

lspA

fkpB

ispH

rihC

dapB

carA

carB

caiF

caiE

caiD

caiC

caiB

caiA

caiT

fixA

fixB

fixC

fixX

yaaU

kefF

kefC

folA

apaH

apaG

rsmA

pdxA

surA

lptD

djlA

rluA

hepA

polB

araD

araA

araB

araC

yabI

thiQ

thiP

tbpA

sgrR

setA

leuD

leuC

leuB

leuA

leuL

leuO

ilvI

ilvH

cra

mraZ

rsmH

ftsL

ftsI

murE

murF

mraY

murD

---

---

Listing Gene Identifiers and Accession Numbers from a BioCyc Genome

This post describes a way to gather the identifiers associated with a gene, which are stored under several different object properties in BioCyc (in some cases referred to as slots).  These identifiers are useful for verifying the identity of gene references between EcoCyc and other gene databases and catalogs.  Use of these identifiers is more reliable than depending on the gene names.

BioCyc PGDBs store identifiers for genes in several places.  These identifiers include the PGDB’s own BioCyc identifier, unification links to other databases (stored as PGDB database links), and locus tags from the Genbank entry for the genome.  These additional identifiers are stored as properties (slots) of the gene frame called Accession-1 and Accession-2.  Different PGDBs may assign different sets of identifiers to these slots, but using these slots allows a consistent way to access these.  In this post, I’ll discuss how to use SmartTables to build a list of identifiers associated with a set of genes.  I’ll use EcoCyc as an example PGDB, both because it uses both accession slots, and because the demonstration doesn’t require an active subscription.  Since it uses SmartTables, you will need a free BioCyc account to follow this demonstration.  Here’s a screenshot of the final table.  The final, full table is also linked here.

 

This is the step-by-step procedure.

1.     Go to EcoCyc.org and login.  If you are already logged in to BioCyc, change your organism to E. coli K12. Substr MG1655.

2.     Now open the ‘Smart Tables’ menu and choose the ‘Special Smart Tables’ command.

3.     This will take you to a page with a list of special smart tables corresponding to many types of entities a BioCyc user may find useful, such as all compounds, genes, or enzymes in E coli.  Click on the “All genes of E. coli...”, which will be the second row in the list.

4.     Since you are logged in, this will create an editable copy of the special SmartTable which lists all the genes in E. coli, including the Gene’s name, and the Accession-1 property, as well the left and right boundaries in the genome and the gene’s product.  However, as I mentioned, there are additional properties with alternative identifiers.

5.     Above the table, there are three drop-down boxes.  The middle one is labeled ‘Add Property Column’.  Additional gene identifiers are available in two columns: Object ID and Accession-2.  Add an Object ID column by clicking on the drop down list and selecting the column by name.  The column will appear at the far right.  This is EcoCyc’s own internal identifier.  Repeat the process for Accession-2.  The Accession-1 and Accession-2 identifiers for E. coli are locus tags from two different naming systems.  As I mentioned, the particular identifiers used will be different in different PGDBs.

6.     You can also add identifiers from one or more external databases in the same way.  Use the Add Property Column dropdown and choose ‘Database Links’.  Now a window with a list of external databases appears.  You can select one, or several while holding the appropriate key (control or command on Macs).  In this example, I selected the EchoBASE database because it has alternate identifiers for many, though not all the genes in EcoCyc.  Click the ‘Go’ button to add the column(s).

7.     Once you have the columns, you can use the right-sidebar Operations Menu on the right-side bar to export the Smart Table to a file.  However, if you’re not interested in saving the coordinates or gene product columns, you can delete those columns by selecting them (click in the colored space immediately above the column name), then choosing the ‘delete column’ command, which appears in both the delete and column submenus in the Operations Menu. 

I hope you have found this discussion useful and I welcome questions and comments.

Local Downloading of BioCyc PGDBs for Pathway Tools Users

BioCyc offers thousands of PGDBS, currently 10,980 to be exact, and while subscribers can access all of these via the BioCyc web site, most desktop Pathway Tools users will want to work with a limited subset of these thousands.

Installing the desktop version of Pathway Tools provides a number of advantages:

• Ability to create and edit PGDBs locally
• Compare your PGDB(s) with similar organisms in BioCyc
• Ability to query PGDB data from Python, Perl, Java, Lisp
• Faster speed
• Develop metabolic models 

Working with a PGDB in desktop Pathway Tools requires having a local copy. Downloading all those nearly 11,000 files would take up a lot of space and downloading time, so we provide two ways to get only the PGDBs you need.

The first download method is to download an appropriate bundle of PGDBs when you download Pathway tools. If you have a BioCyc subscription, you will have a number of options. Without a subscription (if you only have an academic download license), you will only be able to download the bundle that includes MetaCyc and EcoCyc. With a BioCyc subscription, there are seven available bundles, although a few bundle options are not available for all operating systems. Also the bundles for Windows will sometimes contain fewer PGDBs due to space constraints.

Bundle	Contents
EcoCyc and MetaCyc	EcoCyc, MetaCyc
EcoCyc and MetaCyc +BsubCyc+YeastCyc	EcoCyc, MetaCyc, BsubCyc (Bacillus subtilis subtilis 168), Saccharomyces cerevisiae (YeastCyc)
EcoCyc and MetaCyc +Mammals	EcoCyc, MetaCyc, HumanCyc, Mus musculus (MouseCyc)
EcoCyc and MetaCyc  +Tier2	EcoCyc, MetaCyc, BsubCyc, HumanCyc, YeastCyc, Bacillus anthracis Ames (AnthraCyc), Arabodopsis thaliana, Agrobacterium fabrum C58, Caulbacter crescentus CB15, Helicobacter pylori 26695, Leishmania major Friedlin, Mycobacterium tuberculosis H37Rv, M. tuberculosis CDC1551, Peptoclostridium difficile 630, Plasmodium falciparum 3D7, Shigella flexneri 2a str. 2457T (ShigellaCyc), Synechococcus elongatus PC 7942, Vibrio cholera O1 biovar E1 Tor strain N16961
EcoCyc and MetaCyc + Bacilli	EcoCyc, MetaCyc, Anthracyc, BsubCyc, Bacillus amyloliquefaciens DSM 7, B. anthracis Sterne, B. atrophaeus 1942, B. cereus ATCC 14579, B. licheniformis DSM 13, B. megaterium QM B1551, B. pseudofirmus OF4, B. pumilus SAFR-032, B. subtilis spizizenii W23
EcoCyc and MetaCyc + E. coli	EcoCyc, MetaCyc, ShigellaCyc, E. coli 0157:H7 strain EDL933, E. coli CFT073, E. coli K-12 substr. W3110, E. coli UTI89, E. coli O157:H7 str. Sakai, E. coli B str. REL606, E. coli K-12 substr. W3110, E. coli ATCC 8739, E. coli 536, E. coli BL21(DE3), E. coli HS, Salmonella enterica enterica LT2; SGSC 1412; ATCC 700720, Shigella flexneri 301, S. flexneri 8401, S. boydii Sb227, S. sonnei Ss046
EcoCyc and MetaCyc + Mycobacteria	EcoCyc, MetaCyc, Amycolicicoccus subflavus DQS3-9A1, M. avium 104, M. avium paratuberculosis K-10, M. bovis BCG Pasteur 1173P2, M. bovis Tokyo 172, M. gilvum PYR-GCK, M. leprae Br4923, M. marinum M, M. parascrofulaceum ATCC BAA-614, M. sp. MCS MCS, M. sp. JLS JLS, M. sp. KMS, M. gilvum Spyr1, M. sinense JDM601, Mycobacterium tuberculosis H37Rv, M. tuberculosis CDC1551, M. tuberculosis F11, M. tuberculosis KZN 1435, M. ulcerans Agy99, M. vanbaalenii PYR-1

The second download method doesn't limit you to what's available in these pre-built bundles.  SRI maintains an online repository of all SRI created PGDBs as well as dozens of PGDBs submitted by other researchers.  We call this repository the registry.  If you have a BioCyc subscription, you can use the built-in registry import command to download additional PGDBs from within Pathway Tools (Files->Import->Registry).  The importer provides a search capability so you can enter a substring and it will return a list of registry-resident PGDBs whose taxon names match the search string.  Select the taxa you want and go.  Note that after the loading is complete, you will want to close the importer window, which may be hidden by the main pathway tools window.  For more information about the registry, see chapter 6 of the Pathway Tools User's Guide.

We hope you have found this review of ways to get PGDBs helpful.  For most Pathway Tools users, the Registry is the most flexible and frequently the quickest way to set up your collection of PGDBs.