L-leucine catabolism in Pseudomonas fluorescens FW300-N2C3

GapMind for catabolism of small carbon sources

L-leucine catabolism in Pseudomonas fluorescens FW300-N2C3

Best path

livF, livG, livJ, livH, livM, ilvE, bkdA, bkdB, bkdC, lpd, liuA, liuB, liuD, liuC, liuE, atoA, atoD, atoB

Also see fitness data for the top candidates

Rules

Overview: Leucine degradation in GapMind is based on MetaCyc pathway L-leucine degradation I, via branched alpha-keto acid dehydrogenase (link). Other pathways for are not included here because they are not linked to sequence (link) or do not result in carbon incorporation.

all: leucine-transport, ilvE, BKD, liuA, liuB, liuD, liuC, liuE and acetoacetate-degradation

Comment: After transamination to 4-methyl-2-oxopentanoate by ilvE, kbd oxidatively decarboxylates it to isovaleryl-CoA (also known as 3-methylbutanoyl-CoA), liuA oxidizes it to 3-methylcrotonyl-CoA, liuBD carboxylates it to 3-methylglutaconyl-CoA, liuC hydrates it to hydroxymethylglutaryl-CoA, and liuE hydrolyzes it to acetoacetate and acetyl-CoA.

acetoacetate-degradation: acetoacetate-activation and atoB

Comment: The acetoacetate is activated to acetoacetyl-CoA, and cleaved by acetyl-CoA acetyltransferase, giving two acetyl-CoA.

acetoacetate-activation:

atoA and atoD
or aacS
Comment: acetyl-CoA:acetoacetyl-CoA transferase (sometimes given EC 2.8.3.9 or EC 2.8.3.8) or succinyl-CoA:acetoacetyl-CoA transferase (EC 2.8.3.5, also known as 3-oxoacid CoA-transferase) can activate acetoacetate. These have an A and B subunit. Alternatively, an ATP-dependent ligase (aacS) can activate acetoacetate (EC 6.2.1.16).

BKD:

bkdA, bkdB, bkdC and lpd
or vorA, vorB and vorC
or ofoA and ofoB
or ofo
Comment: These decarboxylating dehydrogenases act on 4-methyl-2-oxopentanoate, 3-methyl-2-oxobutanoate (2-oxoisovalerate) and (S)-3-methyl-2-oxopentanoate and are known as the branched-chain alpha-ketoacid dehydrogenases. They can pass electrons to NAD (EC 1.2.1.25) or to ferredoxin (EC 1.2.7.7). The NAD-dependent enzyme is the sum of three activities: EC 1.2.4.4 (the 4-methyl-2-oxopentanoate dehydrogenase, with transfer to the lipopoyllysine residue of 2.3.1.168) which is itself heteromeric, with alpha and beta subunits; EC 2.3.1.168 (dihydrolipoyllysine-residue (3-methylbutanoyl)transferase); and EC 1.8.1.4 (dihydrolipoyl dehydrogenase, transferring electrons to NAD). The well-characterized ferredoxin-dependent enzymes have 3 subunits (vorABC) or 2 subunits (ofoAB). Genetic data identified a fused ferredoxin-dependent enzyme with just 1 subunit (ofo).

leucine-transport:

livF, livG, livJ, livH and livM
or natA, natB, natC, natD and natE
or aapJ, aapQ, aapM and aapP
or leuT
or brnQ
or bcaP
or Bap2
or AAP1
Comment: Transporters were identified using query: transporter:leucine:L-leucine

39 steps (29 with candidates)

Or see definitions of steps

Step	Description	Best candidate	2nd candidate
livF	L-leucine ABC transporter, ATPase component 1 (LivF/BraG)	AO356_05340	AO356_08480
livG	L-leucine ABC transporter, ATPase component 2 (LivG/BraF)	AO356_05335	AO356_08485
livJ	L-leucine ABC transporter, substrate-binding component (LivJ/LivK/BraC/BraC3)	AO356_05320	AO356_08500
livH	L-leucine ABC transporter, permease component 1 (LivH/BraD)	AO356_05325	AO356_08495
livM	L-leucine ABC transporter, permease component 2 (LivM/BraE)	AO356_05330	AO356_08490
ilvE	L-leucine transaminase	AO356_22970	AO356_21330
bkdA	branched-chain alpha-ketoacid dehydrogenase, E1 component alpha subunit	AO356_22990	AO356_24880
bkdB	branched-chain alpha-ketoacid dehydrogenase, E1 component beta subunit	AO356_22985	AO356_24885
bkdC	branched-chain alpha-ketoacid dehydrogenase, E2 component	AO356_22980	AO356_08965
lpd	branched-chain alpha-ketoacid dehydrogenase, E3 component	AO356_22975	AO356_19705
liuA	isovaleryl-CoA dehydrogenase	AO356_01580	AO356_15445
liuB	3-methylcrotonyl-CoA carboxylase, alpha (biotin-containing) subunit	AO356_01595	AO356_02930
liuD	3-methylcrotonyl-CoA carboxylase, beta subunit	AO356_01585	AO356_02920
liuC	3-methylglutaconyl-CoA hydratase	AO356_01590	AO356_24105
liuE	hydroxymethylglutaryl-CoA lyase	AO356_01570	AO356_24105
atoA	acetoacetyl-CoA transferase, A subunit	AO356_21650
atoD	acetoacetyl-CoA transferase, B subunit	AO356_21645
atoB	acetyl-CoA C-acetyltransferase	AO356_21640	AO356_26350
Alternative steps:
aacS	acetoacetyl-CoA synthetase	AO356_00425	AO356_01575
AAP1	L-leucine permease AAP1
aapJ	ABC transporter for amino acids (Asp/Asn/Glu/Pro/Leu), substrate-binding component AapJ	AO356_17380
aapM	ABC transporter for amino acids (Asp/Asn/Glu/Pro/Leu), permease component 2 (AapM)	AO356_17390	AO356_29675
aapP	ABC transporter for amino acids (Asp/Asn/Glu/Pro/Leu), ATPase component AapP	AO356_17395	AO356_18215
aapQ	ABC transporter for amino acids (Asp/Asn/Glu/Pro/Leu), permease component 1 (AapQ)	AO356_17385
Bap2	L-leucine permease Bap2	AO356_17670	AO356_28340
bcaP	L-leucine uptake transporter BcaP
brnQ	L-leucine:Na+ symporter BrnQ/BraB	AO356_19720
leuT	L-leucine:Na+ symporter LeuT
natA	L-leucine ABC transporter, ATPase component 1 (NatA)	AO356_05335	AO356_29020
natB	L-leucine ABC transporter, substrate-binding component NatB
natC	L-leucine ABC transporter, permease component 1 (NatC)	AO356_05330
natD	L-leucine ABC transporter, permease component 2 (NatD)	AO356_08495	AO356_08345
natE	L-leucine ABC transporter, ATPase component 2 (NatE)	AO356_05340	AO356_08480
ofo	branched-chain alpha-ketoacid:ferredoxin oxidoreductase, fused
ofoA	branched-chain alpha-ketoacid:ferredoxin oxidoreductase, alpha subunit OfoA
ofoB	branched-chain alpha-ketoacid:ferredoxin oxidoreductase, beta subunit OfoB
vorA	branched-chain alpha-ketoacid:ferredoxin oxidoreductase, alpha subunit VorA
vorB	branched-chain alpha-ketoacid:ferredoxin oxidoreductase, beta subunit VorB
vorC	branched-chain alpha-ketoacid:ferredoxin oxidoreductase, gamma subunit VorC

Confidence: high confidence medium confidence low confidence
transporter – transporters and PTS systems are shaded because predicting their specificity is particularly challenging.

This GapMind analysis is from Sep 17 2021. The underlying query database was built on Sep 17 2021.

Downloads

Candidates (tab-delimited)
Steps (tab-delimited)
Rules (tab-delimited)
Protein sequences (fasta format)
Organisms (tab-delimited)
SQLite3 databases

Related tools

About GapMind

Each pathway is defined by a set of rules based on individual steps or genes. Candidates for each step are identified by using ublast (a fast alternative to protein BLAST) against a database of manually-curated proteins (most of which are experimentally characterized) or by using HMMer with enzyme models (usually from TIGRFam). Ublast hits may be split across two different proteins.

A candidate for a step is "high confidence" if either:

ublast finds a hit to a characterized protein at above 40% identity and 80% coverage, and bits >= other bits+10.
- (Hits to curated proteins without experimental data as to their function are never considered high confidence.)
HMMer finds a hit with 80% coverage of the model, and either other identity < 40 or other coverage < 0.75.

where "other" refers to the best ublast hit to a sequence that is not annotated as performing this step (and is not "ignored").

Otherwise, a candidate is "medium confidence" if either:

ublast finds a hit at above 40% identity and 70% coverage (ignoring otherBits).
ublast finds a hit at above 30% identity and 80% coverage, and bits >= other bits.
HMMer finds a hit (regardless of coverage or other bits).

Other blast hits with at least 50% coverage are "low confidence."

Steps with no high- or medium-confidence candidates may be considered "gaps." For the typical bacterium that can make all 20 amino acids, there are 1-2 gaps in amino acid biosynthesis pathways. For diverse bacteria and archaea that can utilize a carbon source, there is a complete high-confidence catabolic pathway (including a transporter) just 38% of the time, and there is a complete medium-confidence pathway 63% of the time. Gaps may be due to:

our ignorance of proteins' functions,
omissions in the gene models,
frame-shift errors in the genome sequence, or
the organism lacks the pathway.

GapMind relies on the predicted proteins in the genome and does not search the six-frame translation. In most cases, you can search the six-frame translation by clicking on links to Curated BLAST for each step definition (in the per-step page).

For more information, see:

the paper from 2019 on GapMind for amino acid biosynthesis
the paper from 2022 on GapMind for carbon sources
the source code
instructions for running GapMind on your computer

If you notice any errors or omissions in the step descriptions, or any questionable results, please let us know

by Morgan Price, Arkin group, Lawrence Berkeley National Laboratory

GapMind for catabolism of small carbon sources