Pleiotropic effects of antithrombin strand 1C substitution mutations.

Lane, David A.; Olds, R.J.; Conard, J.; Boisclair, Michael D.; Bock, Susan; Hultin, Mae B.; Abildgaard, Ulrich; Ireland, H; Thompson, Elizabeth A.; Sas, G.

doi:10.1172/jci116133

Cited by 97 publications

(75 citation statements)

References 64 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…A large number of missense ATIII mutations have been discovered in patients suffering from thrombosis (20)(21)(22). To further test the importance of ordered disulfide bond formation and N-linked glycans for in-cell folding, secretion, and function, we monitored the effects of known ATIII mutations that add or subtract Cys residues and one mutation that adds an N-linked glycosylation site using the cellular assays developed in this study (Table S1).…”

Section: Atiiimentioning

confidence: 99%

Cellular folding pathway of a metastable serpin

Chandrasekhar

Wang

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Although proteins generally fold to their thermodynamically most stable state, some metastable proteins populate higher free energy states. Conformational changes from metastable higher free energy states to lower free energy states with greater stability can then generate the work required to perform physiologically important functions. However, how metastable proteins fold to these higher free energy states in the cell and avoid more stable but inactive conformations is poorly understood. The serpin family of metastable protease inhibitors uses large conformational changes that are downhill in free energy to inhibit target proteases by pulling apart the protease active site. The serpin antithrombin III (ATIII) targets thrombin and other proteases involved in blood coagulation, and ATIII misfolding can thus lead to thrombosis and other diseases. ATIII has three disulfide bonds, two near the N terminus and one near the C terminus. Our studies of ATIII in-cell folding reveal a surprising, biased order of disulfide bond formation, with early formation of the C-terminal disulfide, before formation of the N-terminal disulfides, critical for folding to the active, metastable state. Early folding of the predominantly β-sheet ATIII domain in this two-domain protein constrains the reactive center loop (RCL), which contains the proteasebinding site, ensuring that the RCL remains accessible. N-linked glycans and carbohydrate-binding molecular chaperones contribute to the efficient folding and secretion of functional ATIII. The inability of a number of disease-associated ATIII variants to navigate the folding reaction helps to explain their disease phenotypes.

show abstract

Section: Atiiimentioning

confidence: 99%

Cellular folding pathway of a metastable serpin

Chandrasekhar

Wang

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…Congenital AT deficiency is usually heterozygous and classified into two types: quantitative deficiency (type I) and qualitative deficiency (type II). The latter includes reactive site defect, heparin binding site defect, and pleiotrophic effect AT deficiencies [6].…”

Section: Introductionmentioning

confidence: 99%

Molecular basis of antithrombin deficiency in four Japanese patients with antithrombin gene abnormalities including two novel mutations

et al. 2007

View full text Add to dashboard Cite

We analyzed the antithrombin (AT) gene in four unrelated Japanese patients with an AT deficiency, and individually identified four distinct mutations in the heterozygous state. There were two novel mutations, 2417delT leading to a frameshift with a premature termination at amino acid -3 (FS-3Stop) and C2640T resulting in a missense mutation (Ala59Val). Previously reported mutations, T5342C (Ser116Pro) and T72C (Met-32Thr), were also found in the other two patients. To understand the molecular basis responsible for the AT deficiency in these patients, in vitro expression experiments were performed using HEK293 cells transfected with either wild type or respective mutant AT expression vector. We found that -3Stop-AT and -32Thr-AT were not secreted into the culture media, whereas 116Pro-AT and 59Val-AT were secreted normally. We further studied the heparin cofactor activity and the binding to heparin of each recombinant AT molecule. Ser116Pro mutation significantly impaired the binding affinity to heparin resulting in a reduced heparin cofactor activity. In contrast, we found that Ala59Val mutant AT unexpectedly showed a normal affinity to heparin, but severely impaired the heparin cofactor activity. Our findings suggested that FS-3Stop and Met-32Thr mutations are responsible for type I AT deficiency, whereas Ser116Pro and Ala59Val mutations contribute to type II AT deficiency, confirming that there were diverse molecular mechanisms of AT deficiency depend upon discrete AT gene abnormalities as reported previously. Am. J. Hematol. 82:702-705, 2007. V

show abstract

“…22 The mutations reported here generate highmolecular-weight species, probably resulting from misfolding and the formation of intermolecular disulfide bonds with other plasma components. These important structural changes are responsible for the multiple or pleiotropic effects observed.…”

Section: Discussionmentioning

confidence: 83%

Three novel mutations of antithrombin inducing high-molecular-mass compounds.

Emmerich

Vidaud

Alhenc‐Gelas

et al. 1994

Arterioscler Thromb

View full text Add to dashboard Cite

We have identified three novel mutations of the antithrombin (AT) gene in patients with thrombotic complications: a Cys 128 -» Tyr mutation, a G -» A mutation in the intervening sequence 4 (FVS4) 14 nucleotide 5' to exon 5, and a 9 bp deletion in the 3' end of exon 6 resulting in a short aberrant sequence after Arg 425. The latter mutation was associated with an Arg 47 -• His mutation in two compound heterozygous brothers. These three mutations led to the expression in the circulation of small amounts of inactive molecules with a high molecular mass in immunoblot analysis. In reducing conditions, these variant molecules had a normal molecular mass, which led us to postulate that these mutations prevent the formation of one intramolecular disulfidc bond and allow the formation of intermolecular disulfide bonds. Plasma from a heterozygous patient bearing the Cys 128 -• A ntithrombin (AT) is the main natural inhibitor of /\ thrombin and other coagulation proteases, and X A. its physiological importance was first shown in 1965 by the description of a hereditary AT deficiency in a family with unexplained thrombosis.1 The inhibitory effect of AT, due to the almost irreversible trapping of target proteases, is strongly enhanced by natural glycosaminoglycans such as heparin.The AT gene, the nucleotide sequence of which has recently been elucidated, 2 maps more than 13 480 bp, comprises 7 exons (1, 2, 3a, 3b, 4, 5, and 6), and encodes a polypeptide of 464 amino acids. Several intracellular modifications occur before the secretion of the folded protein, such as the formation of three intramolecular disulfide bonds in positions 8-128, 21-95, and 247-430 of the amino-acid sequence, glycosylation at four sites, and cleavage of a 32-amino-acid signal peptide (for a review see Reference 3). The structure of the mature protein presents large homologies with other serine protease inhibitors, forming a superfamily called serpins. Crystallographic study of cleaved a-1-antitrypsin provided a working model of three-dimensional serpin structure, 4 the validity of which was recently confirmed by crystal

show abstract

Pleiotropic effects of antithrombin strand 1C substitution mutations.

Cited by 97 publications

References 64 publications

Cellular folding pathway of a metastable serpin

Cellular folding pathway of a metastable serpin

Molecular basis of antithrombin deficiency in four Japanese patients with antithrombin gene abnormalities including two novel mutations

Three novel mutations of antithrombin inducing high-molecular-mass compounds.

Contact Info

Product

Resources

About