It therefore catalyzes the condensation of 2 molecules of delta-aminolevulinate to form porphobilinogen (a precursor of heme, cytochromes and other hemoproteins). This reaction is the first common step in the biosynthesis of all biological tetrapyrroles. Zinc is essential for enzymatic activity.
The structural basis for allosteric regulation of Porphobilinogen synthase (PBGS) is modulation of a quaternary structure equilibrium between octamer and hexamer (via dimers), which is represented schematically as 6mer* ↔ 2mer* ↔ 2mer ↔ 8mer. The * represents a reorientation between two domains of each subunit that occurs in the dissociated state because it is sterically forbidden in the larger multimers.
PBGS is encoded by a single gene and each PBGS multimer is composed of multiple copies of the same protein. Each PBGS subunit consists of a ~300 residue αβ-barrel domain, which houses the enzyme's active site in its center, and a >25 residue N-terminal arm domain. Allosteric regulation of PBGS can be described in terms of the orientation of the αβ-barrel domain with respect to the N-terminal arm domain.
Each N-terminal arm has up to two interactions with other subunits in a PBGS multimer. One of these interactions helps to stabilize a "closed" conformation of the active site lid. The other interaction restricts solvent access from the other end of the αβ-barrel.
In the inactive multimeric state, the N-terminal arm domain is not involved in the lid-stabilizing interaction, and in the crystal structure of the inactive assembly, the active site lid is disordered.
As a nearly universal enzyme with a highly conserved active site, PBGS would not be a prime target for the development of antimicrobials and/or herbicides. To the contrary, allosteric sites can be much more phylogenetically variable than active sites, thus presenting more drug development opportunities.
Phylogenetic variation in PBGS allostery leads to the framing of discussion of PBGS allosteric regulation in terms of intrinsic and extrinsic factors.
Intrinsic allosteric regulators
The allosteric magnesium ion lies at the highly hydrated interface of two pro-octamer dimers. It appears to be easily dissociable, and it has been shown that hexamers accumulate when magnesium is removed in vitro.
Though it is not common to consider hydronium ions as allosteric regulators, in the case of PBGS, side chain protonation at locations other than the active site has been shown to affect the quaternary structure equilibrium, and thus to affect the rate of its catalyzed reaction as well.
Extrinsic allosteric regulators
Small molecule hexamer stabilization
Inspection of the PBGS 6mer* reveals a surface cavity that is not present in the 8mer. Small molecule binding to this phylogenetically variable cavity has been proposed to stabilize 6mer* of the targeted PBGS and consequently inhibit activity.
Such allosteric regulators are known as morphlocks because they lock PBGS in a specific morpheein form (6mer*).
ALAD enzymatic activity is inhibited by lead, beginning at blood lead levels that were once considered to be safe (<10 μg/dL) and continuing to correlate negatively across the range from 5 to 95 μg/dL. Inhibition of ALAD by lead leads to anemia primarily because it both inhibits heme synthesis and shortens the lifespan of circulating red blood cells, but also by stimulating the excessive production of the hormone erythropoietin, leading to inadequate maturation of red cells from their progenitors. A defect in the ALAD structural gene can cause increased sensitivity to lead poisoning and acute hepatic porphyria. Alternatively spliced transcript variants encoding different isoforms have been identified.
A deficiency of porphobilinogen synthase is usually acquired (rather than herary) and can be caused by heavy metal poisoning, especially lead poisoning, as the enzyme is very susceptible to inhibition by heavy metals.
The morpheein model of allostery exemplified by PBGS adds an additional layer of understanding to potential mechanisms for regulation of protein function and complements the increased focus that the protein science community is placing on protein structure dynamics.
This model illustrates how the dynamics of phenomena such as alternate protein conformations, alternate oligomeric states, and transient protein-protein interactions can be harnessed for allosteric regulation of catalytic activity.
^Beaumont C, Foubert C, Grandchamp B, Weil D, Gross MS, Nordmann Y (May 1984). "Assignment of the human gene for delta aminolevulinate dehydrase to chromosome 9 by somatic cell hybridization and specific enzyme immunoassay". Annals of Human Genetics. 48 (2): 153–9. doi:10.1111/j.1469-1809.1984.tb01010.x. PMID6378062. S2CID24098976.
Doss M, von Tiepermann R, Schneider J (1981). "Acute hepatic porphyria syndrome with porphobilinogen synthase defect". The International Journal of Biochemistry. 12 (5–6): 823–6. doi:10.1016/0020-711X(80)90170-6. PMID7450139.
Kaya AH, Plewinska M, Wong DM, Desnick RJ, Wetmur JG (January 1994). "Human delta-aminolevulinate dehydratase (ALAD) gene: structure and alternative splicing of the erythroid and housekeeping mRNAs". Genomics. 19 (2): 242–8. doi:10.1006/geno.1994.1054. PMID8188255.
Kervinen J, Jaffe EK, Stauffer F, Neier R, Wlodawer A, Zdanov A (July 2001). "Mechanistic basis for suicide inactivation of porphobilinogen synthase by 4,7-dioxosebacic acid, an inhibitor that shows dramatic species selectivity". Biochemistry. 40 (28): 8227–36. CiteSeerX10.1.1.374.9639. doi:10.1021/bi010656k. PMID11444968.