Mistakes GPR rules beta oxidation - ACAD

[ina999111]

The GPRs for mitochondrial acyl-coa dehydrogenase reactions have been discussed in issue #634. Six reactions occurring in the mitochondria were identified to have GPRs with genes involved in peroximal beta-oxidation. The GPRs rules were corrected further than just removing peroximal genes, they were also examined in term of the appropriate ACADs for different chain length. Some guidelines were suggested, most of which I agree with, except for the following:

• In the reference mentioned for ACAD8 (https://doi.org/10.1086/303105), the enzyme is shown to act on 2-methylbutyryl-CoA and isobutyryl-CoA, both of which are short and branched chain fatty acids. The paper states that “None of the enzymes [ACADSB and ACAD8] had significant activity toward either butyryl-CoA or isovaleryl-CoA.” Therefore, I don’t think ACAD8 should be included in the reactions involving butyryl-CoA and propanoyl-CoA (MAR03163 and MAR03212 respectively)
• For ACADS, considering the scale of the axis of the figure referenced (https://doi.org/10.1016/j.ymgme.2010.12.005), I would argue that this enzyme can act on C4-C6.
• The reference mentioned for ACAD9 (https://doi.org/10.1074/jbc.M504460200) shows that the enzyme has a high activity toward C9 to C11, low activity toward C12 and high activity toward C14 to C22. However, this pattern seems odd and is discordant with another study that shows activity toward C14 to 20 (https://doi.org/10.1016/j.ymgme.2010.12.005). Unless there is a biological explanation why ACAD9 would act on C10 and C14 but not C12, I would trust the pattern found in the second paper.

Otherwise, these rules also match my own findings from reading literature on ACAD enzymes substrate specificity.

Enzyme	Can handle unsaturation?	Chain lengths (issue #634)	Chain lengths (my suggestion)
ACADVL	yes	12-24	12-24
ACADL	yes	6-24	6-24
ACADM	no	6-16	6-16
ACADS	no	4	4-6
ACADSB	no	<=6	<=6
ACAD8	no	3-4	Branched only
ACAD9	yes	9-11, 14-22	14-20
ACAD11	unknown	20-24	20-24

Many reactions do not follow these rules. Some “lumped” reaction (describing a whole beta-oxidation cycle or multiple cycles) have GPR rules with only 1 or multiple ACADs. This is incorrect since a beta-oxidation cycle consist of at least 4 different reactions (more for unsaturated acyl-CoA’s). I will not discuss these since my understanding from other issues is that these lumped reactions are getting removed in favor or higher resolution reactions.

Here is a list of the reactions describing the first step of the beta-oxidation cycle (excluding branched fatty acids), with the current GPR and my suggested GPR based on the table above:

ID	formula_readable	formula fatty acid	current gpr	suggested gpr
MAR03212	FAD + propanoyl-CoA --> acrylyl-CoA + FADH2	C3:0	ACADM or ACADS or ACAD8 or ACAD9 or ACADSB or ACAD11	ACADS or ACADSB
MAR03163	butyryl-CoA + FAD --> crotonyl-CoA + FADH2	C4:0	ACADM or ACADS or ACAD8 or ACAD9 or ACADSB or ACAD11	ACADS or ACADSB
MAR03444	arachidonyl-CoA + FAD --> 2-trans-cis,cis,cis,cis-4,8,11,14-eicosapentaenoyl-CoA + FADH2	C20:4(5,8,11,14)	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03448	dihomo-gamma-linolenoyl-CoA + FAD --> FADH2 + trans-2-cis,cis,cis-8,11,14-eicosatetraenoyl-CoA	C20:3(8,11,14)	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03452	FAD + gamma-linolenoyl-CoA --> (2E,6Z,9Z,12Z)-octadecatetraenoyl-CoA + FADH2	C18:3(6,9,12)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03107	docosanoyl-CoA + FAD --> (2E)-docosenoyl-CoA + FADH2	C22:0	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03111	eicosanoyl-CoA + FAD --> (2E)-eicosenoyl-CoA + FADH2	C20:0	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03115	FAD + stearoyl-CoA --> (2E)-octadecenoyl-CoA + FADH2	C18:0	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03121	FAD + palmitoyl-CoA --> (2E)-hexadecenoyl-CoA + FADH2	C16:0	ACADVL or ACADL or ACADM or ACAD9	ok as is
MAR03128	FAD + myristoyl-CoA --> (2E)-tetradecenoyl-CoA + FADH2	C14:0	ACADVL or ACOX3 or ACADL or ACADM or ACOX1 or ACAD9	ACADVL or ACADL or ACADM or ACAD9
MAR03135	FAD + lauroyl-CoA --> (2E)-dodecenoyl-CoA + FADH2	C12:0	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACADM
MAR03142	decanoyl-CoA + FAD --> (2E)-decenoyl-CoA + FADH2	C10:0	ACOX3 or ACOX1 or ACADM	ACADL or ACADM
MAR03149	FAD + octanoyl-CoA --> (2E)-octenoyl-CoA + FADH2	C8:0	ACOX3 or ACOX1 or ACADM or ACAD9	ACADL or ACADM
MAR03156	FAD + hexanoyl-CoA --> (2E)-hexenoyl-CoA + FADH2	C6:0	ACOX3 or ACOX1 or ACADSB	ACADL or ACADM or ACASB
MAR03170	FAD + heneicosanoyl-CoA --> (2E)-heneicosenoyl-CoA + FADH2	C21:0	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03174	FAD + nonadecanoyl-CoA --> (2E)-nonadecenoyl-CoA + FADH2	C19:0	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03178	FAD + heptadecanoyl-CoA --> (2E)-heptadecenoyl-CoA + FADH2	C17:0	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03182	FAD + pentadecanoyl-CoA --> (2E)-pentadecenoyl-CoA + FADH2	C15:0	ACADVL or ACADL or ACADM or ACAD9	ok as is
MAR03186	FAD + tridecanoyl-CoA --> (2E)-tridecenoyl-CoA + FADH2	C13:0	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACADM
MAR03190	FAD + undecanoyl-CoA --> (2E)-undecenoyl-CoA + FADH2	C11:0	ACADM or ACAD9	ACADL or ACADM
MAR03194	FAD + nonanoyl-CoA --> (2E)-nonenoyl-CoA + FADH2	C9:0	ACADM or ACAD9	ACADL or ACADM
MAR03198	FAD + heptanoyl-CoA --> (2E)-heptenoyl-CoA + FADH2	C7:0	ACADM or ACAD9	ACADL or ACADM or ACADSB
MAR03202	FAD + pentanoyl-CoA --> (2E)-pentenoyl-CoA + FADH2	C5:0	ACADSB	ACADSB or ACADS
MAR03240	(13Z)-eicosenoyl-CoA + FAD --> FADH2 + trans,cis-2,13-eicosadienoyl-CoA	20:1(13)	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03244	cis-vaccenoyl-CoA + FAD --> FADH2 + trans,cis-octadeca-2,11-dienoyl-CoA	C18:1(11)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03250	FAD + palmitoleoyl-CoA --> FADH2 + trans,cis-hexadeca-2,9-dienoyl-CoA	C16:1(9)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03258	(7Z)-tetradecenoyl-CoA + FAD --> FADH2 + trans,cis-myristo-2,7-dienoyl-CoA	C14:1(7)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03218	(13Z)-docosenoyl-CoA + FAD --> FADH2 + trans,cis-2,13-docosadienoyl-CoA	C22:1(13)	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03222	(11Z)-eicosenoyl-CoA + FAD --> FADH2 + trans,cis-2,11-eicosadienoyl-CoA	C20:1(11)	ACADVL or ACADL or ACADM or ACAD9	ACADL or ACADVL or ACAD11 or ACAD9
MAR03226	FAD + oleoyl-CoA --> FADH2 + trans,cis-octadeca-2,9-dienoyl-CoA	C18:1(9)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03230	7-hexadecenoyl-CoA + FAD --> FADH2 + trans,cis-hexadeca-2,7-dienoyl-CoA	C16:1(7)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03234	5-tetradecenoyl-CoA + FAD --> FADH2 + trans,cis-myristo-2,5-dienoyl-CoA	C14:1(5)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03275	FAD + linoleoyl-CoA --> FADH2 + trans,cis,cis-2,9,12-octadecatrienoyl-CoA	C18:2(9,12)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03280	cis,cis-palmito-7,10-dienoyl-CoA + FAD --> FADH2 + trans,cis,cis-2,7,10-hexadecatrienoyl-CoA	C16:2(7,10)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03284	cis,cis-myristo-5,8-dienoyl-CoA + FAD --> FADH2 + trans-2-cis,cis-5,8-tetradecatrienoyl-CoA	C14:2(5,8)	ACADVL or ACADL or ACADM or ACAD9	ACADVL or ACADL or ACAD9
MAR03294	4-cis-decenoyl-CoA + FAD --> 2-trans-4-cis-decadienoyl-CoA + FADH2	C10:1(4)	ACADVL or ACADL or ACADM or ACAD9	ACADL
MAR03291	FADH2 + Trans,Cis-Dodeca-2,5-Dienoyl Coenzyme A <=> (5Z)-dodecenoyl-CoA + FAD	C12:2(2,5)	ACADL or ACADM or ACOX1 or ACADSB	ACADVL or ACADL
MAR04978	FAD + 4,7-Decadienoyl Coenzyme A --> FADH2 + 2,4,7-Decatrienoyl Coenzyme A	C10:2(4,7)	ACADM	ACADL
MAR05298	FAD + 4,7,10,13-Hexadecatetraenoyl Coenzyme A --> FADH2 + 2,4,7,10,13-Hexadecapentaenoyl Coenzyme A	C16:4(4,7,10,13)	ACADVL	ACADVL or ACADL or ACAD9
MAR05380	(4Z,7Z,10Z,13Z,16Z)-docosapentaenoyl-CoA + FAD --> FADH2 + 2,4,7,10,13,16-Docosahexenoyl Coenzyme A	C25:5(4,7,10,13,16)	ACADVL	ACADL or ACADVL or ACAD11 or ACAD9
MAR05408	(4Z,7Z,10Z,13Z,16Z,19Z)-docosahexaenoyl-CoA + FAD --> FADH2 + 2,4,7,10,13,16,19-Docosaheptenoyl Coenzyme A	C26:6(4,7,10,13,16,19)	ACADVL	ACADL or ACADVL or ACAD11 or ACAD9
MAR06378	FAD + Trans4Decenoyl Coenzyme A --> FADH2 + Cis2Trans4Decadienoyl Coenzyme A	C10:1(4)	ACADM	ACADL
MAR11744	FAD + heptadecanoyl-CoA --> FADH2 + Trans-Delta-2-Heptadecanoyl Coenzyme A	C17:0	ACADL	ACADVL or ACADL or ACAD9

[JHL-452b]

It's really nice result! The mitochondrial acetyl-CoA dehydrogenation process is a very complex part of human metabolism. Although we have done a lot of curation work before, it seems that there are still many problems left. I have checked the location information of all genes and found no errors. I also agree with your conclusion about gene function. There is basically no problem for me. Maybe we need someone more professional to review it.

@Devlin-Moyer Would you be able to review this issue? Apologies if you're busy!

[Devlin-Moyer]

I appreciate the thorough double-checking of my attempts to clean up fatty acid metabolism, since it's a mess and super easy to miss things. I checked to see if any of the reactions you proposed new GPRs for here were also mentioned in any of the other open issues I have about fatty acid oxidation reactions, and found a few overlaps:

• In Remove peroxisomal genes from mitochondrial ACAD reactions #943, I proposed that we change the GPRs for MAR03212 and MAR03163 to ACADS or ACADSB or ACAD8, so I'm gonna go edit that now to remove ACAD8 from those, since you are definitely right that the paper I cited about ACAD8 clearly says that it is not active against butyryl-CoA
• Also in Remove peroxisomal genes from mitochondrial ACAD reactions #943, I proposed to add ACAD11 to the GPRs of MAR03107 and MAR03111, but you've also proposed to remove ACADM. I will also update that pull request to remove ACADM from those GPRs, since I agree with your rationale -- I missed that earlier because I was only trying to ensure that those changes wouldn't leave ACAD11 associated with no reactions, so I never checked to see if MAR03107 and MAR03111 had any other issues
• In Bad GPRs for Beta-Oxidation of Linoleoyl- and gamma-Linolenoyl-CoA Reactions #762, I proposed that ACADVL be removed from the GPRs of MAR03275, MAR03280, and MAR03284, but you've proposed that we keep it in those GPRs. I double checked the paper that I cited in that issue when I said that ACADVL is not active against unsaturated acyl-CoAs, and realized that it actually says that ACADVL is not active with unsaturated acyl-CoAs with 4,5 or 5,6 double bonds, but is active with unsaturated acyl-CoAs with double bonds that are further away. So I agree with your GPRs for MAR03275 (linoleoyl-CoA, double bonds at carbons 9 and 12) and MAR03280 (cis,cis-palmito-7,10-dienoyl-CoA) and will update my issue to reflect that. However, I stand by my earlier claim that ACADVL should not be in the GPR of MAR03284 (cis,cis-myristo-5,8-dienoyl-CoA), and also think that ACADVL should not be in the GPRs of MAR03444, MAR03234, MAR03291, MAR05298, MAR05380, or MAR05480 (but agree with the rest of the genes you have in those GPRs)

Aside from the minor changes I mentioned above, I think all of the rest of your GPRs look correct.