Cobalamin biosynthesis

{{Main|Vitamin B12|l1=Vitamin B₁₂}}

Cobalamin (vitamin B₁₂) is the largest and most structurally complex vitamin. It consists of a modified tetrapyrrole, a corrin, with a centrally chelated cobalt ion and is usually found in one of two biologically active forms: methylcobalamin and adenosylcobalamin. Most prokaryotes, as well as animals, have cobalamin-dependent enzymes that use it as a cofactor, whereas plants and fungi do not use it. In bacteria and archaea, these enzymes include methionine synthase, ribonucleotide reductase, glutamate and methylmalonyl-CoA mutases, ethanolamine ammonia-lyase, and diol dehydratase.{{cite journal |doi=10.1074/jbc.M305837200 |doi-access = free |title=Comparative Genomics of the Vitamin B12 Metabolism and Regulation in Prokaryotes |year=2003 |last1=Rodionov |first1=Dmitry A. |last2=Vitreschak |first2=Alexey G. |last3=Mironov |first3=Andrey A. |last4=Gelfand |first4=Mikhail S. |journal=Journal of Biological Chemistry |volume=278 |issue=42 |pages=41148–41159 |pmid=12869542 }} In certain mammals, cobalamin is obtained through the diet, and is required for methionine synthase and methylmalonyl-CoA mutase.{{cite journal |doi=10.1021/cb6001174 |title=B12 Trafficking in Mammals: A Case for Coenzyme Escort Service |year=2006 |last1=Banerjee |first1=Ruma |journal=ACS Chemical Biology |volume=1 |issue=3 |pages=149–159 |pmid=17163662 }} In humans, it plays essential roles in folate metabolism and in the synthesis of the citric acid cycle intermediate, succinyl-CoA.{{cite web |url=http://lpi.oregonstate.edu/mic/vitamins/vitamin-B12|title=Vitamin B12 |publisher=Micronutrient Information Center, Linus Pauling Institute, Oregon State University, Corvallis, OR|date=4 June 2015 |access-date=20 April 2020}}

There are at least two distinct cobalamin biosynthetic pathways in bacteria:{{cite book |doi=10.1016/s0083-6729(01)61009-4 |chapter=Multiple biosynthetic pathways for vitamin B12: Variations on a central theme |title=Cofactor Biosynthesis |series=Vitamins & Hormones |year=2001 |last1=Roessner |first1=Charles A. |last2=Santander |first2=Patricio J. |last3=Scott |first3=A.Ian |volume=61 |pages=267–297 |pmid=11153269 |isbn=9780127098616 }}

• Aerobic pathway that requires oxygen and in which cobalt is inserted late in the pathway;{{cite journal |doi=10.1042/BST0330815 |title=Aerobic synthesis of vitamin B12: Ring contraction and cobalt chelation |year=2005 |last1=Heldt |first1=D. |last2=Lawrence |first2=A.D. |last3=Lindenmeyer |first3=M. |last4=Deery |first4=E. |last5=Heathcote |first5=P. |last6=Rigby |first6=S.E. |last7=Warren |first7=M.J. |s2cid=37362827 |journal=Biochemical Society Transactions |volume=33 |issue=4 |pages=815–819 |pmid=16042605 }}{{cite web |url=https://biocyc.org/META/NEW-IMAGE?type=PATHWAY&object=P381-PWY |title=Pathway: adenosylcobalamin biosynthesis II (aerobic) |author=R. Caspi |publisher=MetaCyc Metabolic Pathway Database |date=2013-09-25 |access-date=2020-04-24 }} found in Pseudomonas denitrificans and Rhodobacter capsulatus.
• Anaerobic pathway in which cobalt insertion is the first committed step towards cobalamin synthesis;{{cite journal |vauthors=Roessner CA, Huang KX, Warren MJ, Raux E, Scott AI | title = Isolation and characterization of 14 additional genes specifying the anaerobic biosynthesis of cobalamin (vitamin B12) in Propionibacterium freudenreichii (P. shermanii) | journal = Microbiology | volume = 148 | issue = Pt 6 | pages = 1845–1853 |date=June 2002 | pmid = 12055304 | doi = 10.1099/00221287-148-6-1845 | doi-access = free }}{{cite journal |doi=10.1042/BST0330811 |title=Anaerobic synthesis of vitamin B12: Characterization of the early steps in the pathway |year=2005 |last1=Frank |first1=S. |last2=Brindley |first2=A.A. |last3=Deery |first3=E. |last4=Heathcote |first4=P. |last5=Lawrence |first5=A.D. |last6=Leech |first6=H.K. |last7=Pickersgill |first7=R.W. |last8=Warren |first8=M.J. |journal=Biochemical Society Transactions |volume=33 |issue=4 |pages=811–814 |pmid=16042604 }}{{cite web |url=https://biocyc.org/META/NEW-IMAGE?type=PATHWAY&object=PWY-5507 |title=Pathway: adenosylcobalamin biosynthesis I (anaerobic) |author=R. Caspi |publisher=MetaCyc Metabolic Pathway Database |date=2013-09-25 |access-date=2020-04-24 }} found in Salmonella typhimurium, Bacillus megaterium, and Propionibacterium freudenreichii subsp. shermanii.

Either pathway can be divided into two parts:

• Corrin ring synthesis leading to cobyrinic acid, with seven carboxylate groups. In the anaerobic pathway this already contains cobalt but in the aerobic pathway the material formed at that stage is hydrogenobyrinic acid, without the bound cobalt.{{cite journal| title=How Nature builds the pigments of life |last = Battersby | first = A. R. | journal =Pure and Applied Chemistry | year =1993 | volume = 65 |issue = 6 | pages =1113–1122 |url=https://www.degruyter.com/downloadpdf/j/pac.1993.65.issue-6/pac199365061113/pac199365061113.pdf |doi=10.1351/pac199365061113|s2cid = 83942303 }}{{cite journal| title=Tetrapyrroles: the Pigments of Life. A Millennium review |last = Battersby | first = A. R. | journal =Nat. Prod. Rep. | year =2000 | volume = 17 |issue = 6 | pages =507–526 |doi=10.1039/B002635M | pmid = 11152419}}{{cite journal |last1=Fang |first1=H |last2=Kang |first2=J |last3=Zhang |first3=D |title=Microbial production of vitamin B₁₂: a review and future perspectives. |journal=Microbial Cell Factories |date=30 January 2017 |volume=16 |issue=1 |pages=15 |doi=10.1186/s12934-017-0631-y |pmid=28137297 |pmc=5282855 |doi-access=free }}
• Insertion of cobalt, where not already present; formation of amides on all but one of the carboxylate groups to give cobyric acid; attachment of an adenosyl group as ligand to the cobalt; attachment of an aminopropanol sidechain to the one free carboxylic group and assembly of the nucleotide loop which will provide the second ligand for the cobalt.{{cite journal |vauthors=Raux E, Schubert HL, Warren MJ | title = Biosynthesis of cobalamin (vitamin B12): a bacterial conundrum | journal = Cell. Mol. Life Sci. | volume = 57 | issue = 13–14 | pages = 1880–1893 |date=December 2000 | pmid = 11215515 | doi = 10.1007/PL00000670| s2cid = 583311 | pmc = 11147154 }}

A further type of synthesis occurs through a salvage pathway, where outside corrinoids are absorbed to make B₁₂.

Species from the following genera and the following individual species are known to synthesize cobalamin: Propionibacterium shermanii, Pseudomonas denitrificans, Streptomyces griseus, Acetobacterium, Aerobacter, Agrobacterium, Alcaligenes, Azotobacter, Bacillus, Clostridium, Corynebacterium, Flavobacterium, Lactobacillus, Micromonospora, Mycobacterium, Nocardia, Proteus,

Rhizobium, Salmonella, Serratia, Streptococcus and Xanthomonas.{{cite journal|vauthors=Perlman D|year=1959|title=Microbial synthesis of cobamides|journal=Advances in Applied Microbiology|volume=1|pages=87–122|doi=10.1016/S0065-2164(08)70476-3|isbn=9780120026012|pmid=13854292}}{{cite journal|vauthors=Martens JH, Barg H, Warren MJ, Jahn D|date=March 2002|title=Microbial production of vitamin B12|journal=Applied Microbiology and Biotechnology|volume=58|issue=3|pages=275–285|doi=10.1007/s00253-001-0902-7|pmid=11935176|s2cid=22232461}}

{{Main|Porphyrin#Synthesis}}

In the early steps of the biosynthesis, a tetrapyrrolic structural framework is created by the enzymes deaminase and cosynthetase which transform aminolevulinic acid via porphobilinogen and hydroxymethylbilane to uroporphyrinogen III. The latter is the first macrocyclic intermediate common to haem, chlorophyll, sirohaem and cobalamin itself.{{cite journal | vauthors = Battersby AR, Fookes CJ, Matcham GW, McDonald E | title = Biosynthesis of the pigments of life: formation of the macrocycle | journal = Nature | volume = 285 | issue = 5759 | pages = 17–21 | date = May 1980 | pmid = 6769048 | doi = 10.1038/285017a0 | bibcode = 1980Natur.285...17B | s2cid = 9070849 | doi-access = free }}{{cite journal | vauthors = Frank S, Brindley AA, Deery E, Heathcote P, Lawrence AD, Leech HK, Pickersgill RW, Warren MJ | display-authors = 6 | title = Anaerobic synthesis of vitamin B12: characterization of the early steps in the pathway | journal = Biochemical Society Transactions | volume = 33 | issue = Pt 4 | pages = 811–814 | date = August 2005 | pmid = 16042604 | doi = 10.1042/BST0330811 }}

The biosynthesis of cobalamin diverges from that of haem and chlorophyll at uroporphrinogen III: its transformation involves the sequential addition of methyl (CH₃) groups to give intermediates that were given trivial names according to the number of these groups that have been incorporated. Hence, the first intermediate is precorrin-1, the next is precorrin-2 and so on. The incorporation of all eight additional methyl groups which occur in cobyric acid was investigated using ¹³C methyl-labelled S-adenosyl methionine. It was not until scientists at Rhône-Poulenc Rorer used a genetically-engineered strain of Pseudomonas denitrificans, in which eight of the cob genes involved in the biosynthesis of the vitamin had been overexpressed, that the complete sequence of methylation and other steps could be determined, thus fully establishing all the intermediates in the pathway.{{cite journal |doi=10.1128/jb.175.22.7430-7440.1993 |title=Biosynthesis of the corrin macrocycle of coenzyme B12 in Pseudomonas denitrificans |year=1993 |last1=Debussche |first1=L. |last2=Thibaut |first2=D. |last3=Cameron |first3=B. |last4=Crouzet |first4=J. |last5=Blanche |first5=F. |journal=Journal of Bacteriology |volume=175 |issue=22 |pages=7430–7440 |pmid=8226690 |pmc=206888 }}{{Cite book |title=The 1702 chair of chemistry at Cambridge: transformation and change|editor-first1=Mary D. |editor-last1=Archer |editor-first2= Christopher D. |editor-last2=Haley |last = Battersby | first = Alan |chapter= Chapter 11: Discovering the wonder of how Nature builds its molecules |pages=xvi, 257–282 |publisher=Cambridge University Press |year=2005 |isbn=0521828732}}

The enzyme CobA catalyses two methylations, to give precorrin-2:{{cite journal |doi=10.1042/bj2650725 |title=The Escherichia coli cysG gene encodes S-adenosylmethionine-dependent uroporphyrinogen III methylase |year=1990 |last1=Warren |first1=M. J. |last2=Roessner |first2=C. A. |last3=Santander |first3=P. J. |last4=Scott |first4=A. I. |journal=Biochemical Journal |volume=265 |issue=3 |pages=725–729 |pmid=2407234 |pmc=1133693 }}

:File:Dihydrosirochlorin.png

:(1a) uroporphyrinogen III + S-adenosyl methionine $\backslash rightleftharpoons$ precorrin-1 + S-adenosyl-L-homocysteine

:(1b) precorrin-1 + S-adenosyl methionine $\backslash rightleftharpoons$ precorrin-2 + S-adenosyl-L-homocysteine

The enzyme CobI then converts this to precorrin-3A:

:precorrin-2 + S-adenosyl methionine $\backslash rightleftharpoons$ precorrin-3A + S-adenosyl-L-homocysteine

Next, the enzyme CobG transforms precorrin-3A to precorrin-3B:

:precorrin-3A + NADH + H⁺ + O₂ $\backslash rightleftharpoons$ precorrin-3B + NAD⁺ + H₂O

This enzyme is an oxidoreductase that requires oxygen and hence the reaction can only operate under aerobic conditions. The naming of these precorrins as 3A and 3B reflects the fact that each contains three more methyl groups than uroporphyrinogen III but with different structures: in particular, precorrin-3B has an internal γ-lactone ring formed from the ring A acetic acid sidechain closing back on to the macrocycle.

The enzyme CobJ continues the theme of methyl group insertion. Importantly, during this step the macrocycle ring-contracts so that the product contains for the first time the corrin core which characterises cobalamin.

:File:Precorrin-3B to Precorrin-4.svg

:precorrin-3B + S-adenosyl methionine $\backslash rightleftharpoons$ precorrin-4 + S-adenosyl-L-homocysteine

Methyl group insertions continue as the enzyme CobM acts on precorrin-4:{{cite journal |doi=10.1039/b108967f |title=The biosynthesis of adenosylcobalamin (Vitamin B12) |year=2002 |last1=Warren |first1=Martin J. |last2=Raux |first2=Evelyne |last3=Schubert |first3=Heidi L. |last4=Escalante-Semerena |first4=Jorge C. |journal=Natural Product Reports |volume=19 |issue=4 |pages=390–412 |pmid=12195810 }}

:precorrin-4 + S-adenosyl methionine $\backslash rightleftharpoons$ precorrin-5 + S-adenosyl-L-homocysteine

The newly-inserted methyl group is added to ring C at the carbon attached to the methylene (CH₂) bridge to ring B. This is not its final location on cobalamin as a later step involves its rearrangement to an adjacent ring carbon.

The enzyme CobF now removes the acetyl group located at position 1 of the ring system in precorrin-4 and replaces it with a newly-introduced methyl group. The name of the product, precorrin-6A, reflects the fact that six methyl groups in total have been added to uroporphyrinogen III up to this point. However, since one of these has been extruded with the acetate group, the structure of precorrin-6A contains just the remaining five.

:File:Precorrin-5 to precorrin-6A.svg

:precorrin-5 + S-adenosyl methionine + H₂O $\backslash rightleftharpoons$ precorrin-6A + S-adenosyl-L-homocysteine + acetate

The enzyme CobK now reduces a double bond in ring D using NADPH:

:precorrin-6A + NADPH + H⁺ $\backslash rightleftharpoons$ precorrin-6B + NADP⁺

Precorrin-6B therefore differs in structure from precorrin-6A only by having an extra two hydrogen atoms.

The enzyme CobL has two active sites, one catalysing two methyl group additions and the other the decarboxylation of the CH₂COOH group on ring D, so that this substituent becomes a simple methyl group:

:File:Precorrin-6B to precorrin-8.svg

:precorrin-6B + 2 S-adenosyl methionine $\backslash rightleftharpoons$ precorrin-8X + 2 S-adenosyl-L-homocysteine + CO₂

The enzyme CobH catalyzes a rearrangement reaction, with the result that the methyl group that had been added to ring C is isomerised to its final location, an example of intramolecular transfer:{{cite journal |doi=10.1128/jb.174.3.1043-1049.1992 |title=The final step in the biosynthesis of hydrogenobyrinic acid is catalyzed by the cobH gene product with precorrin-8x as the substrate |year=1992 |last1=Thibaut |first1=D. |last2=Couder |first2=M. |last3=Famechon |first3=A. |last4=Debussche |first4=L. |last5=Cameron |first5=B. |last6=Crouzet |first6=J. |last7=Blanche |first7=F. |journal=Journal of Bacteriology |volume=174 |issue=3 |pages=1043–1049 |pmid=1732194 |pmc=206186 }}

:precorrin-8X $\backslash rightleftharpoons$ hydrogenobyrinate

The next enzyme in the pathway, CobB, selectively converts two of the eight carboxylic acid groups into their primary amides. ATP is used to provide the energy for amide bond formation, with the transferred ammonia coming from glutamine:{{cite journal |doi=10.1128/jb.172.11.6239-6244.1990 |title=Purification and characterization of cobyrinic acid a,c-diamide synthase from Pseudomonas denitrificans |year=1990 |last1=Debussche |first1=L. |last2=Thibaut |first2=D. |last3=Cameron |first3=B. |last4=Crouzet |first4=J. |last5=Blanche |first5=F. |journal=Journal of Bacteriology |volume=172 |issue=11 |pages=6239–6244 |pmid=2172209 |pmc=526805 }}

:hydrogenobyrinic acid + 2 ATP + 2 glutamine + 2 H₂O $\backslash rightleftharpoons$ hydrogenobyrinic acid a,c-diamide + 2 ADP + 2 phosphate + 2 glutamic acid

Cobalt(II) insertion into the macrocycle is catalysed by the enzyme Cobalt chelatase (CobNST):{{cite journal |doi=10.1128/JB.174.22.7445-7451.1992 |title=Assay, purification, and characterization of cobaltochelatase, a unique complex enzyme catalyzing cobalt insertion in hydrogenobyrinic acid a,c-diamide during coenzyme B12 biosynthesis in Pseudomonas denitrificans |year=1992 |last1=Debussche |first1=L. |last2=Couder |first2=M. |last3=Thibaut |first3=D. |last4=Cameron |first4=B. |last5=Crouzet |first5=J. |last6=Blanche |first6=F. |journal=Journal of Bacteriology |volume=174 |issue=22 |pages=7445–7451 |pmid=1429466 |pmc=207441 }}

:hydrogenobyrinic acid a,c-diamide + Co²⁺ + ATP + H₂O $\backslash rightleftharpoons$ {{Not a typo|cob(II)yrinic}} acid a,c-diamide + ADP + phosphate + H⁺

It is at this stage that the aerobic pathway and the anaerobic pathway merge, with later steps being chemically identical.

Many of the steps beyond uroporphyrinogen III in anaerobic organisms such as Bacillus megaterium involve chemically similar but genetically distinct transformations to those in the aerobic pathway.{{cite journal |doi=10.1128/JB.00918-06 |title=Fine-Tuning Our Knowledge of the Anaerobic Route to Cobalamin (Vitamin B12) |year=2006 |last1=Roessner |first1=Charles A. |last2=Scott |first2=A. Ian |journal=Journal of Bacteriology |volume=188 |issue=21 |pages=7331–7334 |pmid=16936030 |pmc=1636268 }}

The key difference in the pathways is that cobalt is inserted early in anaerobic organisms by first oxidising precorrin-2 to its fully aromatised form sirohydrochlorin and then to that compound's cobalt(II) complex.{{cite journal |doi=10.1042/BST20120066 |title=The anaerobic biosynthesis of vitamin B12 |year=2012 |last1=Moore |first1=Simon J. |last2=Warren |first2=Martin J. |journal=Biochemical Society Transactions |volume=40 |issue=3 |pages=581–586 |pmid=22616870 |s2cid=26057998 }} These reactions are catalysed by CysG and Sirohydrochlorin cobaltochelatase.{{cite journal |doi=10.1007/s10969-006-9008-x |title=Crystal Structure of the Vitamin B12 Biosynthetic Cobaltochelatase, CbiXS, from Archaeoglobus Fulgidus |year=2006 |last1=Yin |first1=Jiang |last2=Xu |first2=Linda X. |last3=Cherney |first3=Maia M. |last4=Raux-Deery |first4=Evelyne |last5=Bindley |first5=Amanda A. |last6=Savchenko |first6=Alexei |last7=Walker |first7=John R. |last8=Cuff |first8=Marianne E. |last9=Warren |first9=Martin J. |last10=James |first10=Michael N. G. |journal=Journal of Structural and Functional Genomics |volume=7 |issue=1 |pages=37–50 |pmid=16835730 |s2cid=6613060 }}

As in the aerobic pathway, the third methyl group is introduced by a methyltransferase enzyme, CbiL:

:cobalt-sirohydrochlorin + S-adenosyl methionine $\backslash rightleftharpoons$ cobalt-factor III + S-adenosyl-L-homocysteine

Methylation and ring contraction to form the corrin macrocycle occurs next, catalysed by the enzyme Cobalt-factor III methyltransferase (CbiH, {{EC number|2.1.1.272}}){{cite journal |doi=10.1074/jbc.M112.422535 |title=Characterization of the Enzyme CbiH60Involved in Anaerobic Ring Contraction of the Cobalamin (Vitamin B12) Biosynthetic Pathway |year=2013 |last1=Moore |first1=Simon J. |last2=Biedendieck |first2=Rebekka |last3=Lawrence |first3=Andrew D. |last4=Deery |first4=Evelyne |last5=Howard |first5=Mark J. |last6=Rigby |first6=Stephen E. J. |last7=Warren |first7=Martin J. |journal=Journal of Biological Chemistry |volume=288 |issue=1 |pages=297–305 |pmid=23155054 |pmc=3537027 |doi-access=free }}

:File:Cobalt-precorrin-3 to cobalt-precorrin-4.svg

:cobalt-factor III + S-adenosyl methionine $\backslash rightleftharpoons$ cobalt-precorrin-4 + S-adenosyl-L-homocysteine

In this pathway, the resulting material contains a δ-lactone, a six-membered ring, rather than the γ-lactone (five-membered ring) of precorrin-3B.

The introduction of the methyl group at C-11 in the next step is catalysed by Cobalt-precorrin-4 methyltransferase (CbiF, {{EC number|2.1.1.271}}){{cite journal |doi=10.1021/ja062940a |title=Genetically Engineered Synthesis and Structural Characterization of Cobalt−Precorrin 5A and −5B, Two New Intermediates on the Anaerobic Pathway to Vitamin B12: Definition of the Roles of the CbiF and CbiG Enzymes |year=2006 |last1=Kajiwara |first1=Yasuhiro |last2=Santander |first2=Patricio J. |last3=Roessner |first3=Charles A. |last4=Pérez |first4=Lisa M. |last5=Scott |first5=A. Ian |journal=Journal of the American Chemical Society |volume=128 |issue=30 |pages=9971–9978 |pmid=16866557 |bibcode=2006JAChS.128.9971K }}

:cobalt-precorrin-4 + S-adenosyl methionine $\backslash rightleftharpoons$ cobalt-precorrin-5 + S-adenosyl-L-homocysteine

The scene is now set for the extrusion of the two-carbon fragment corresponding to the acetate released in the formation of precorrin-6A in the aerobic pathway. In this case the fragment released is acetaldehyde and this is catalysed by CbiG:

: cobalt-precorrin-5A + H₂O $\backslash rightleftharpoons$ cobalt-precorrin-5B + acetaldehyde + 2 H⁺

The steps from cobalt-precorrin-5B to {{Not a typo|cob(II)yrinic}} acid a,c-diamide in the anaerobic pathway are essentially chemically identical to those in the aerobic sequence. The intermediates are called cobalt-precorrin-6A, cobalt-precorrin-6B, cobalt-precorrin-8 and cobyrinic acid. The enzymes in sequence are CbiD;{{cite journal | doi = 10.1074/jbc.M501805200 | doi-access = free | title = Genetically Engineered Production of 1-Desmethylcobyrinic Acid, 1-Desmethylcobyrinic Acida,c-Diamide, and Cobyrinic Acida,c-Diamide in Escherichia coli Implies a Role for CbiD in C-1 Methylation in the Anaerobic Pathway to Cobalamin | year = 2005 | last1 = Roessner | first1 = Charles A. | last2 = Williams | first2 = Howard J. | last3 = Scott | first3 = A. Ian | journal = Journal of Biological Chemistry | volume = 280 | issue = 17 | pages = 16748–16753 | pmid = 15741157 }} Cobalt-precorrin-6A reductase (CbiJ, {{EC number|1.3.1.106}});{{cite journal |doi=10.1155/2005/903614 |title=Role of the precorrin 6-X reductase gene in cobamide biosynthesis in Methanococcus maripaludis |year=2005 |last1=Kim |first1=Wonduck |last2=Major |first2=Tiffany A. |last3=Whitman |first3=William B. |journal=Archaea |volume=1 |issue=6 |pages=375–384 |pmid=16243778 |pmc=2685584 |doi-access=free }} CbiT, Cobalt-precorrin-8 methylmutase (CbiC, {{EC number|5.4.99.60}}) and CbiA. The final enzyme forms {{Not a typo|cob(II)yrinic}} acid a,c-diamide as the two pathways converge.

Aerobic and anaerobic organisms share the same chemical pathway beyond {{Not a typo|cob(II)yrinic}} acid a,c-diamide and this is illustrated for the cob gene products.

The cobalt(II) is reduced to {{Not a typo|cobalt(I)}} by the enzyme {{Not a typo and then the enzyme {{Not a typo attaches an adenosyl ligand to the metal. Next, the enzyme CobQ converts all the carboxylic acids, except the propionic acid on ring D, to their primary amides.

In aerobic organisms, the enzyme CobCD now attaches (R)-1-amino-2-propanol (derived from threonine) to the propionic acid, forming adenosylcobinamide and the enzyme CobU phosphorylates the terminal hydroxy group to form adenosylcobinamide phosphate. The same final product is formed in anaerobic organisms by direct reaction of adenosylcobyric acid with (R)-1-amino-2-propanol O-2-phosphate (derived from threonine-O-phosphate by the enzyme CobD) catalysed by the enzyme CbiB.

In a separate branch of the pathway, 5,6-dimethylbenzimidazole is biosynthesised from flavin mononucleotide by the enzyme 5,6-dimethylbenzimidazole synthase and converted by CobT to alpha-ribazole 5' phosphate. Then the enzyme CobU activates adenosylcobinamide phosphate by formation of adenosylcobinamide-GDP and CobV links the two substrates to form Adenosylcobalamin-5'-phosphate. In the final step to the coenzyme, CobC removes the 5' phosphate group:{{cite web |url=https://biocyc.org/META/NEW-IMAGE?type=PATHWAY&object=PWY-5509 |title=Pathway: adenosylcobalamin biosynthesis from adenosylcobinamide-GDP I|author=R. Caspi |publisher=MetaCyc Metabolic Pathway Database |date=2007-04-23 |access-date=2020-04-24 }}{{cite journal |doi=10.1128/jb.01665-06 |title=Reassessment of the Late Steps of Coenzyme B12 Synthesis in Salmonella enterica: Evidence that Dephosphorylation of Adenosylcobalamin-5′-Phosphate by the CobC Phosphatase is the Last Step of the Pathway |year=2007 |last1=Zayas |first1=Carmen L. |last2=Escalante-Semerena |first2=Jorge C. |journal=Journal of Bacteriology |volume=189 |issue=6 |pages=2210–2218 |pmid=17209023 |pmc=1899380 }}

:Adenosylcobalamin-5'-phosphate + H₂O $\backslash rightleftharpoons$ adenosylcobalamin + phosphate

The complete biosynthetic route involves a long linear path that requires about 25 contributing enzyme steps.

Many prokaryotic species cannot biosynthesize adenosylcobalamin, but can make it from cobalamin. These organisms are capable of cobalamin transport into the cell and its conversion to the required coenzyme form.{{cite web |url=https://biocyc.org/META/NEW-IMAGE?type=PATHWAY&object=PWY-6268|title=Pathway: adenosylcobalamin salvage from cobalamin |author=R. Caspi |publisher=MetaCyc Metabolic Pathway Database |date=2013-09-25 |access-date=2020-04-24 }} Even organisms such as Salmonella typhimurium that can make cobalamin also assimilate it from external sources when available.{{cite journal |doi=10.1128/jb.172.1.273-280.1990 |title=CobA function is required for both de novo cobalamin biosynthesis and assimilation of exogenous corrinoids in Salmonella typhimurium |year=1990 |last1=Escalante-Semerena |first1=J. C. |last2=Suh |first2=S. J. |last3=Roth |first3=J. R. |journal=Journal of Bacteriology |volume=172 |issue=1 |pages=273–280 |pmid=2403541 |pmc=208428 }}{{cite journal |doi=10.1128/jb.185.24.7193-7201.2003 |title=A New Pathway for Salvaging the CoenzymeB12 Precursor Cobinamide in Archaea Requires Cobinamide-Phosphate Synthase (CbiB) Enzyme Activity |year=2003 |last1=Woodson |first1=Jesse D. |last2=Zayas |first2=Carmen L. |last3=Escalante-Semerena |first3=Jorge C. |journal=Journal of Bacteriology |volume=185 |issue=24 |pages=7193–7201 |pmid=14645280 |pmc=296239 }}{{cite journal |doi=10.1073/pnas.0305939101 |title=CbiZ, an amidohydrolase enzyme required for salvaging the coenzyme B12 precursor cobinamide in archaea |year=2004 |last1=Woodson |first1=J. D. |last2=Escalante-Semerena |first2=J. C. |journal=Proceedings of the National Academy of Sciences |volume=101 |issue=10 |pages=3591–3596 |pmid=14990804 |pmc=373507 |bibcode=2004PNAS..101.3591W |doi-access=free }} Uptake into cells is facilitated by ABC transporters which absorb the cobalamin through the cell membrane.{{cite journal |doi=10.1128/JB.187.17.5901-5909.2005 |title=ABC Transporter for Corrinoids in Halobacterium sp. Strain NRC-1 |year=2005 |last1=Woodson |first1=Jesse D. |last2=Reynolds |first2=April A. |last3=Escalante-Semerena |first3=Jorge C. |journal=Journal of Bacteriology |volume=187 |issue=17 |pages=5901–5909 |pmid=16109931 |pmc=1196138 }}

{{Main|MMACHC}}

In humans, dietary sources of cobalamin are bound after ingestion as transcobalamins.{{cite web |url=https://biocyc.org/META/NEW-IMAGE?type=PATHWAY&object=PWY-7974 |title=Pathway: cobalamin salvage (eukaryotic) |author=R. Caspi |publisher=MetaCyc Metabolic Pathway Database |date=2013-09-25 |access-date=2020-04-24 }} They are then converted to the coenzyme forms in which they are used. Methylmalonic aciduria and homocystinuria type C protein is the enzyme which catalyzes the decyanation of cyanocobalamin as well as the dealkylation of alkylcobalamins including methylcobalamin and adenosylcobalamin.{{cite journal |doi=10.1016/j.ymgme.2009.04.005 |title=Processing of alkylcobalamins in mammalian cells: A role for the MMACHC (CBLC) gene product |year=2009 |last1=Hannibal |first1=Luciana |last2=Kim |first2=Jihoe |last3=Brasch |first3=Nicola E. |last4=Wang |first4=Sihe |last5=Rosenblatt |first5=David S. |last6=Banerjee |first6=Ruma |last7=Jacobsen |first7=Donald W. |journal=Molecular Genetics and Metabolism |volume=97 |issue=4 |pages=260–266 |pmid=19447654 |pmc=2709701 }}{{cite journal |doi=10.1016/j.cbpa.2009.07.007 |title=The tinker, tailor, soldier in intracellular B12 trafficking |year=2009 |last1=Banerjee |first1=Ruma |last2=Gherasim |first2=Carmen |last3=Padovani |first3=Dominique |journal=Current Opinion in Chemical Biology |volume=13 |issue=4 |pages=484–491 |pmid=19665918 |pmc=5750051 }}{{cite journal |doi=10.1111/j.1365-2141.2009.07937.x |title=Advances in the understanding of cobalamin assimilation and metabolism |year=2010 |last1=Quadros |first1=Edward V. |journal=British Journal of Haematology |volume=148 |issue=2 |pages=195–204 |pmid=19832808 |pmc=2809139 }}

• {{cite book |doi= 10.1016/B978-008045382-8.00144-1 |chapter= Biosynthesis of Heme and Vitamin B12 |title= Comprehensive Natural Products II |year= 2010 |last1= Layer |first1= Gunhild |last2= Jahn |first2= Dieter |last3= Deery |first3= Evelyne |last4= Lawrence |first4= Andrew D. |last5= Warren |first5= Martin J. |pages= 445–499 |isbn= 9780080453828 }}

{{Reflist}}

• [https://www.youtube.com/watch?v=TGMs3S_B8Wg Prof Sir Alan Battersby: the biosynthesis of Vitamin B₁₂] St. Catharine's College, Cambridge, video

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Category:Protein families

Category:Vitamin B12

Category:Biosynthesis