As text, or see rules and steps
# Methionine biosynthesis in GapMind is based on MetaCyc pathways # L-methionine biosynthesis I via O-succinylhomoserine and cystathionine (metacyc:HOMOSER-METSYN-PWY), # II via O-phosphohomoserine and cystathionine (metacyc:PWY-702), # III via O-acetylhomoserine (metacyc:HSERMETANA-PWY), # or IV with reductive sulfhydrylation of aspartate semialdehyde (metacyc:PWY-7977). # These pathways vary in how aspartate semialdehyde is reduced and sulfhydrylated to homocysteine. # GapMind does not represent the formation of the methyl donors for methionine synthase, # such as 5-methyltetrahydrofolate or methyl corrinoid proteins. # For BRENDA::O63067 -- the paper describes a monofunctional hom but the sequence of uniprot:O63067 is # much longer and has a close homolog of functional aspartate kinase (due to alternative splicing?). # In Corynebacterium, aspartate kinase has two subunits, both apparently encoded by the same gene by # using start codons (PMID:1956296); Q46133 is the shorter regulatory subunit and lacks the catalytic domain, # so it does not suffice for activity and is ignored. asp-kinase aspartate kinase EC:2.7.2.4 ignore:BRENDA::O63067 ignore:BRENDA::Q46133 asd aspartate semi-aldehyde dehydrogenase EC:1.2.1.11 # Ga0059261_2711 (uniprot:A0A1L6J6Q3) from Sphingomonas koreensis DSMZ 15582 # is distant from characterized homoserine dehydrogenases # and is confirmed by fitness data. # BT2403 from Bacteroides thetaiotaomicron (uniprot:Q8A541_BACTN) # is a fusion of aspartate kinase and homoserine dehydrogenase. # The homoserine dehydrogenase portion is somewhat diverged. Its role is confirmed by strong cofitness with # threonine synthase (the defined media for B. thetaiotaomicron included methionine). # DvMF_1412 from Desulfovibrio vulgaris Miyazaki F (uniprot:B8DRS3_DESVM) is a somewhat # diverged homoserine dehydrogenase; it has auxotrophic phenotypes. # uniprot:A0A168MV81 from Brevundimonas is a somewhat diverged homoserine dehydrogenase (fused to aspartate kinase); # it has auxotrophic phenotypes. hom homoserine dehydrogenase EC:1.1.1.3 uniprot:A0A1L6J6Q3 uniprot:Q8A541_BACTN uniprot:B8DRS3_DESVM uniprot:A0A168MV81 # As discussed in PMID:28581482, many members of the MetA family are # actually homoserine O-acetyltransferases, and many members of the MetX family # are actually homoserine O-succinyltransferases. Fortunately, many # enzymes of both types have been curated in Swiss-Prot. metA homoserine O-succinyltransferase EC:2.3.1.46 # MetX is often encoded next to a methyltransferase-like protein MetW. # Because MetW is not consistently required for MetX's activity, it is # not included in GapMind. MetW can bind MetX and increase its # activity (PMID:33604638), and in some bacteria, there is tight # cofitness between MetX and MetW, which suggests that MetW is # required for MetX's activity (i.e., Paraburkholderia bryophila, many # Pseudomonas, Caulobacter crescentus, or Sphingomonas koreensis). # But in other diverse bacteria, metW mutants can still grow in minimal media # (i.e., Herbaspirillum seropedicae or Cupriavidus necator), or have # milder phenotypes than metX mutants (i.e., Burkholderia # phytofirmans, Acidovorax 3H11, Dechlorosoma suillum PS, Marinobacter # adhaerens). metX homoserine O-acetyltransferase EC:2.3.1.31 hom_kinase homoserine kinase EC:2.7.1.39 # Many metB proteins have some activity as O-succinylhomoserine sulfhydrylase (metZ) as well, # and many metZ proteins are annotated with this EC number. So this step # may include both enzymes. # METI_BACSU (uniprot:O31631) has activity as CGS but is given a more vague EC number. # uniprot:Q1M0P5 is now thought to be a gamma-lyase instead (see metacyc:HP_RS00540-MONOMER), so it # is ignored. metB cystathionine gamma-synthase EC:2.5.1.48 curated:SwissProt::O31631 ignore:SwissProt::Q1M0P5 # BRENDA annotates Q84UD0 with this EC number but it may be a cystine lyase only (PMID:12525491). # BRENDA annotates Q9EYM7 with this EC but it appears to be a cystathionine beta synthase (PMID:12101301). # uniprot:Q9SIV0 appears to be specific to glucosinolate biosynthesis. # CH_088676 is annotated as this but with a different EC number, so it is ignored. # uniprot:Q4606 is annotated as cysteine desulfidase but is nearly identical to uniprot:Q93QC6, which is # a cystathionine beta-lyase, so it is ignored. # XoMetC (uniprot:Q5H4T8) is annotated as a gamma-lyase by BRENDA, but it also has beta-lyase activity # (PMID:24531493). metC cystathionine beta-lyase EC:4.4.1.13 ignore:BRENDA::Q84UD0 ignore:BRENDA::Q9EYM7 ignore:SwissProt::Q9SIV0 ignore:CharProtDB::CH_088676 ignore:BRENDA::Q46061 curated:BRENDA::Q5H4T8 # METI_BACSU (uniprot:O31631) has activity as OAS but is given a more vague EC number. # CH_123612, uniprot:Q9WZY4, and uniprot:Q5SK88 are annotated as this but without the EC number. metY O-acetylhomoserine sulfhydrylase EC:2.5.1.49 curated:SwissProt::O31631 ignore_other:O-succinylhomoserine sulfhydrylase curated:CharProtDB::CH_123612 curated:SwissProt::Q9WZY4 curated:SwissProt::Q5SK88 # No EC number for metZ, so use "O-succinylhomoserine sulfhydrylase", which matches # uniprot:METZ_PSEAE and uniprot:METZ_MYCTU. metZ O-succinylhomoserine sulfhydrylase term:O-succinylhomoserine sulfhydrylase ignore_other:EC 2.5.1.49 metE vitamin B12-independent methionine synthase EC:2.1.1.14 # In many thermophilic archaea, MetE seems to be split into two pieces (PMC7857596). There # is experimental support for a protein complex (PMC2668238), and the two pieces # often appear to form an operon, but there is no experimental evidence that they # are a methionine synthase. We added the gene from Haloferax volcanii (uniprot:D4GW95) as it is # diverged but is conserved next to a catalytic component with correct functional residues (uniprot:D4GW90). split_metE_1 vitamin B12-independent methionine synthase, folate-binding component predicted:G0EDA1_PYRF1 predicted:D4GW95 # In many thermophilic archaea, MetE seems to be split into two pieces (see PMC7857596). # The catalytic component has the necessary zinc-binding residues (H219, C221, C307) # and homocysteine-binding residues (S20, E71, D185). # We added the gene from Haloferax volcanii (uniprot:D4GW90) as it # has the correct functional residues and is conserved next to a potential folate-binding component # (uniprot:D4GW90). split_metE_2 vitamin B12-independent methionine synthase, catalytic component predicted:G0EFB7_PYRF1 predicted:D4GW90 # Desulfovibrio have a somewhat diverged MetH, without the activation domain, but confirmed by # cofitness (DVU1585 = uniprot:Q72BP9_DESVH is cofit with MetF; DvMF_0476 = uniprot:B8DKK4_DESVM is cofit with a RamA- # like activation protein). # 3-part split MetH proteins from Phaeobacter are ignored. metH vitamin B12-dependent methionine synthase EC:2.1.1.13 ignore:reanno::Phaeo:GFF1501 ignore:reanno::Phaeo:GFF1318 ignore:reanno::Phaeo:GFF1321 ignore:reanno::Phaeo:GFF1319 ignore:reanno::Phaeo:GFF1582 uniprot:Q72BP9_DESVH uniprot:B8DKK4_DESVM # In Phaeobacter and some related bacteria, MetH is split into 3 parts (PMC5764234) split_metH_1 Methionine synthase component, B12 binding and B12-binding cap domains curated:reanno::Phaeo:GFF1319 split_metH_2 Methionine synthase component, methyltransferase domain curated:reanno::Phaeo:GFF1321 split_metH_3 Methionine synthase component, pterin-binding domain curated:reanno::Phaeo:GFF1582 # In E. coli and many other bacteria, the MetH protein includes a reactivation domain (pfam:PF02965), # but other ATP-dependent (ramA-like) activation proteins are also thought to exist. # Ignore MetH proteins, as they often contain the reactivation domain and this # creates confusion when checking for reverse hits. # In Heliobacterium modesticaldum, the missing reactivation domain is probably provided # by H1S01_RS06050 (very similar to A0A6I3SQJ4), which does not hit the HMM but for PF02965 but is found by PFam-N # or foldseek, and is usually encoded to next to MetH. B12-reactivation-domain MetH reactivation domain hmm:PF02965 ignore_other:EC 2.1.1.13 predicted:A0A6I3SQJ4 # As of April 2019, all characterized members of the RamA family or PF14574 are involved in the reactivation # of co(II)balamin. This includes RamA (uniprot:B8Y445), DvMF_1398, PGA1_c15200, and ELI_0370 (part of a O-demethylase). # Many bacteria contain MetH and probably rely on a distant homolog of RamA for reactivation of B-12. # pfam:PF14574 describes only the C-terminal putative ATPase domain of RamA, but no other functions are known, # except for the reactivation of Co(II) corrinoid proteins (i.e. RamQ, uniprot:P0DX10). ramA ATP-dependent reduction of co(II)balamin hmm:PF14574 term:ATP-dependent reduction of co(II)balamin # In the reductive sulfuration of aspartate semialdehyde, the # sulfurtransferase component is # MA1821 or DvMF_1464 (see PMID:25315403 and PMC5764234) or uniprot:Q57564 (from SwissProt). # Although this reaction has not been biochemically demonstrated, a distant homolog performs a similar # reaction, converting sulfoacetaldehyde to coenzyme M (see MJ1681 and PMID:30932481). # (This family was formerly DUF39.) asd-S-transferase sulfuration of L-aspartate semialdehyde, persulfide component curated:SwissProt::Q8TPT4 curated:reanno::Miya:8500721 curated:SwissProt::Q57564 # The NIL/ferredoxin component is MA1822 or DvMF_0262 (see PMID:25315403 and PMC5764234). # In Methanococcus, the persulfide component (MEVAN_RS03425, uniprot:A6UQ02) is diverged but # is in a conserved operon with the sulfurtransferase component. asd-S-ferredoxin reductive sulfuration of L-aspartate semialdehyde, ferredoxin component curated:SwissProt::Q8TPT3 curated:reanno::Miya:8499492 predicted:A6UQ02 # The putative persulfide forming component is MA1715 or DvMF_0044 (see PMID:25315403 and PMC5764234). # A conserved cysetien in MA1821 is modified to a persulfide in vivo (PMID:28165724). # This component is not 100% required in Methanosarcina acetivorans (possible redundancy). # In Hippea alviniae, this protein (G415_RS0107280, similar to uniprot:F2LX84) is diverged but is in an operon with the other components. asd-S-perS putative persulfide forming protein uniprot:Y1715_METAC curated:reanno::Miya:8499265 predicted:F2LX84 # Methanogens have a short homolog of MetE that transfers methyl groups from methylcobalamin # (not 5-methyltetrahydrofolates) to homocysteine to form methionine (PMID:10469143). # We named this family of "core" methioine synthases MesA and proposed # that MtrA (the corrinoid subunit of methyltetrahydromethanopterin:coenzyme M methyltransferase) # is the physiological methyl donor (PMC7857596). mesA Methylcobalamin:homocysteine methyltransferase MesA curated:SwissProt::P55299 # Another core methionine synthase (distantly related to MesA) has been characterized # in Dehalococcoides (PMC7005905). # It probably obtains methyl groups from the iron-sulfur corrinoid protein of the # Wood-Ljungdahl pathway (CoFeSP), but this is not proven. # We named this family MesB (PMID:33534785). mesB Methylcobalamin:homocysteine methyltransferase MesB curated:metacyc::MONOMER-21502 # MesC is another family of core methionine synthases, without experimental evidence, but # with the correct functional residues, and linked to the Wood-Ljungdahl pathway # by the gene neighbor method # (PMC7857596). The corrinoid protein of the Wood-Ljungdahl pathway is probably the methyl donor. mesC Methylcobalamin:homocysteine methyltransferase MesC predicted:Q8TUL3_METAC # Genetic evidence shows that ACIAD3523 and Ga0059261_2929 are methionine synthases, # see PMC2290942 and PMID:33534785. # They require mesX (ACIAD3524 or Ga0059261_2928) and oxygen for activity, but not # 5-methyltetrahydrofolates or cobalamin. mesD oxygen-dependent methionine synthase, methyltransferase component MesD uniprot:Q6F6Z8 curated:reanno::Korea:Ga0059261_2929 # MesX is required for the activity of MesD, see PMC2290942. mesX oxygen-dependent methionine synthase, putative oxygenase component MesX uniprot:Q6F6Z7 curated:reanno::Korea:Ga0059261_2928 aspartate-semialdehyde: asp-kinase asd # Reductive sulfhydrylation of aspartate semialdehyde to homocysteine is carried out by # a multi-component system (see PMID:25315403 and PMC5764234) asd-sulfhydrylation: asd-S-transferase asd-S-ferredoxin asd-S-perS homoserine: aspartate-semialdehyde hom # Transsulfuration is the conversion of homoserine to homocysteine, with the sulfur being obtained from cysteine. # It is thought to occur with any # of the activated forms of homoserine (O-acetyl-, O-succinyl-, or O-phospho-homoserine). transsulfuration: metA metB metC transsulfuration: metX metB metC transsulfuration: hom_kinase metB metC # Homocysteine can be formed by reduction of aspartate semialdehyde, direct sulfurylation of activated homoserine, # or transsulfuration of (activated) homoserine. # Activated forms of homoserine include O-acetylhomoserine, O-succinylhomoserine, or O-phospho-homoserine. homocysteine: aspartate-semialdehyde asd-sulfhydrylation homocysteine: homoserine metX metY homocysteine: homoserine metA metZ homocysteine: homoserine transsulfuration # MetH occasionally oxidizes the vitamin B12 cofactor from Co(I) to Co(II), so # a reductase is needed to maintain its activity. B12-reactivation: B12-reactivation-domain B12-reactivation: ramA # Besides MetH (with B-12 reactivation) or 3-part MetH as in Phaeobacter (PMC5764234), or MetE, # or MetE split into two parts (PMC7857596), # GapMind also includes the folate-independent systems MesA, MesB, # MesC, and MesD/MesX (PMC7857596). # It is possible that the corrinoid-dependent methionine synthases (MesA, MesB, or MesC) would require B12 reactivation, # but this is not proven, and some methanogens with MesA seem to lack RamA, so # B12 reactivation is not included. # The role of split MetE or MesC lacks experimental evidence, and is based on # gene neighborhoods and functional residues only (PMC7857596). methionine_synthase: metH B12-reactivation methionine_synthase: split_metH_1 split_metH_2 split_metH_3 B12-reactivation methionine_synthase: metE methionine_synthase: split_metE_1 split_metE_2 methionine_synthase: mesA methionine_synthase: mesB methionine_synthase: mesC methionine_synthase: mesD mesX all: homocysteine methionine_synthase
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:
Otherwise, a candidate is "medium confidence" if either:
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:
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:
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