Pathogenesis

 Site specificity of conformational mutations

A defining characteristic of conformational diseases is that they arise from mutations that result in a disadvantageous gain of function. Such mutations have consequences over and above any accompanying loss of function.

So, the liver cirrhosis that is associated with alpha1-antitrypsin deficiency arises from conformationally destabilizing mutations of alpha1-antitrypsin, but not from the null mutations that result in total non-expression.

 Similarly, whereas non-expressing null variants of antithrombin result in a predisposition to thrombosis, an even greater threat of thrombosis occurs with conformationally destabilizing variants.

These carry with them, as well as plasma deficiency, the risk of pathological monomeric and polymeric conformational transitions that inactivate the protein, with an accompanying sudden and severe onset of thrombosis.

 Another characteristic of conformationally destabilizing mutations is their site specificity. This is seen in the serpins: some 50 such mutations are specifically located in the key mobile regions of these molecules - the hinges of the reactive loop, and the shutter region that underlies the focal opening of the beta-sheet A.

The prime example of a hinge mutation is seen with the replacement, in Z alpha1-antitrypsin, of the conserved glutamate of the proximal hinge of the reactive loop by lysine. The special significance of this replacement, in conformational terms, has been borne out by the subsequent identification of the identical mutation in different situations.

The first was in a family with a deficiency of the plasma anticoagulant heparin cofactor II ; the second was the mutation in Drosophila that is responsible for the serpin dysfunction that produces constitutive activation of antifungal peptides and epidermal necrosis - the necrotic (Nec) phenotype.

 However, the most frequent site of destabilizing mutations in the serpins is in the shutter region, which is formed by the closely packed amino-acid side-chains on which the sliding movement of the strands of the beta-sheet A takes place.

The significance of the shutter region, and in particular of its highly conserved core sequence (amino-acid residues 51-56 on the serpin template numbering), was shown by the finding that two other mutants of alpha1-antitrypsin were associated with plasma deficiency and hepatic inclusions: alpha1-antitrypsin Siiyama (S53F)36 and alpha1-antitrypsin Mmalton (52F deleted).

The Siiyama mutation is the most common cause of severe alpha1-antitrypsin deficiency in Japan and the Mmalton (also known as Mnichinan and Mcagliari) variant is the commonest cause of severe alpha1-antitrypsin deficiency in Sardinia.

Both mutations cause polymerization and the retention of 1-antitrypsin in hepatocytes. Similar mutations in the shutter region of antithrombin (P54T, P54S, S56N) favour the spontaneous formation of polymers and the retention of antithrombin in hepatocytes, with a consequently increased risk of thrombosis.

Other shutter mutations in C1 inhibitor (F52S, P54L) and alpha1-antichymotrypsin (L55P) also cause polymerization and the retention of protein in the liver.

The reduction in the levels of circulating protein in individuals with deficiencies in C1q inhibitor and alpha1-antichymotrypsin allows the uncontrolled activity of proteinases in the complement and inflammatory cascades and hence the clinical syndromes of angio-oedema ANGIO-OEDEMA and panlobar emphysema.

 The other vulnerable site for mutations in the serpins is the distal hinge of the reactive-centre loop. Numerous mutations here, especially in antithrombin and C1 inhibitor9, result in a thermally sensitive instability that is seen clinically as episodes of thrombosis or angio-oedema triggered by incidental infection and fever.

As well as these main regions of vulnerability, mutations in less crucial regions of the molecule can also predispose to polymer formation, as with the S (E264V) and I (R39C) variants of 1-antitrypsin. The point mutations that are responsible for these variants have less effect on beta-sheet A than does the Z variant.

So, the rates of polymer formation are much slower than for Z alpha1-antitrypsin, and results in less retention of protein in hepatocytes, milder plasma deficiency and the lack of a clinical phenotype.

However, if a mild, slowly polymerizing S or I variant of alpha1-antitrypsin is inherited with a rapidly polymerizing Z variant, then the two can interact to form heteropolymeric inclusions in hepatocytes and associated cirrhosis.

 Propagation and prions

Structural studies of the serpins have also shown the feasibility of another perplexing feature of some of the familial and acquired encephalopathies: the ability of the underlying protein oligomers and filaments to self-propagate and even, with the prions, to propagate infectively.

Aggregation to form polymeric filaments is particularly likely to occur when there is a beta-strand receptor, as in the main beta-sheet of the conformationally unstable variants of alpha1-antitrypsin.

Both the serpins and the prions readily dimerize by domain swapping, and the initial oligomers, like those formed by serpins, act as a template for propagation of the conformational change.

The change is propagated from molecule to molecule, which then extend to give long-chain polymers. This is well illustrated by mutations of antithrombin.

Whereas mutations in alpha1-antitrypsin and neuroserpin result in the formation of long-chain polymers, those in antithrombin result in the formation of an inactive antithrombin monomer.

Moreover, in a process similar to that proposed for the prion encephalopathies, this aberrant form of antithrombin then binds to a normal antithrombin molecule, which leads to the propagation of conformational inactivation.

A more direct insight into the mechanisms of prion propagation that underlie the spongiform encephalopathies is provided by studies of the unrelated yeast prion Ure2. This normally soluble and highly ordered molecule undergoes a conformational change to form fibrils that bind the dye Congo red and show typical amyloid-like birefringence.

However, these fibrils do not have the cross-beta-structure of amyloid and, as is also observed with fibrils formed by orderly aggregation of serpins, the component molecules of the fibril retain their helical structure.

These amyloid-like fibrils are self-propagating but lose this ability over time, coincident with a transition of the fibril to give the typical amyloid X-ray fibre diffraction pattern.

These findings fit with a range of others and together they indicate that the neuronal pathology of the conformational dementias results from the early stages of intracellular aggregation, before or incidental to amyloid formation.

See also

 proteases inhibitors (PIs)
 serpins
 conformational diseases

References

 Lomas DA, Carrell RW. Serpinopathies and the conformational dementias. Nat Rev Genet. 2002 Oct;3(10):759-68. PMID: #12360234#