{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,9,8]],"date-time":"2026-09-08T14:02:21Z","timestamp":1788876141853,"version":"build-2803163510"},"posted":{"date-parts":[[2023,11,2]]},"group-title":"Genetics","reference-count":52,"publisher":"openRxiv","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2023,11,2]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>AlphaMissense is a recently developed method that is designed to classify missense variants into pathogenic, benign, or ambiguous categories across the entire human proteome. Asparagine Synthetase Deficiency (ASNSD) is a developmental disorder associated with severe symptoms, including congenital microcephaly, seizures, and premature death. Diagnosing ASNSD relies on identifying mutations in the asparagine synthetase (ASNS) gene through DNA sequencing and determining whether these variants are pathogenic or benign. Pathogenic ASNS variants are predicted to disrupt the protein\u2019s structure and\/or function, leading to asparagine depletion within cells and inhibition of cell growth. AlphaMissense offers a promising solution for the rapid classification of ASNS variants established by DNA sequencing and provides a community resource of pathogenicity scores and classifications for newly diagnosed ASNSD patients. Here, we assessed AlphaMissense\u2019s utility in ASNSD by benchmarking it against known critical residues in ASNS and evaluating its performance against a list of previously reported ASNSD-associated variants. We also present a pipeline to calculate AlphaMissense scores for any protein in the UniProt database. AlphaMissense accurately attributed a high average pathogenicity score to known critical residues within the two ASNS active sites and the connecting intramolecular tunnel. The program successfully categorized 78.9% of known ASNSD-associated missense variants as pathogenic. The remaining variants were primarily labeled as ambiguous, with a smaller proportion classified as benign. This study underscores the potential role of AlphaMissense in classifying ASNS variants in suspected cases of ASNSD, potentially providing clarity to patients and their families grappling with ongoing diagnostic uncertainty.<\/jats:p>","DOI":"10.1101\/2023.10.30.564808","type":"posted-content","created":{"date-parts":[[2023,11,2]],"date-time":"2023-11-02T13:45:18Z","timestamp":1698932718000},"source":"Crossref","is-referenced-by-count":10,"title":["Utility of AlphaMissense predictions in Asparagine Synthetase deficiency variant classification"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-1746-6320","authenticated-orcid":false,"given":"Stephen J.","family":"Staklinski","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2230-2013","authenticated-orcid":false,"given":"Armin","family":"Scheben","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3557-7219","authenticated-orcid":false,"given":"Adam","family":"Siepel","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Michael S.","family":"Kilberg","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2023110623051071000_2023.10.30.564808v1.1","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2013.08.013"},{"key":"2023110623051071000_2023.10.30.564808v1.2","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.R117.819060"},{"key":"2023110623051071000_2023.10.30.564808v1.3","doi-asserted-by":"crossref","first-page":"97","DOI":"10.1016\/j.gene.2019.04.024","article-title":"Molecular diagnosis of asparagine synthetase (ASNS) deficiency in two Indian families and literature review of 29 ASNS deficient cases","volume":"704","year":"2019","journal-title":"Gene"},{"key":"2023110623051071000_2023.10.30.564808v1.4","doi-asserted-by":"crossref","first-page":"331","DOI":"10.1016\/j.cca.2022.04.989","article-title":"An intractable epilepsy phenotype of ASNS novel mutation in two patients with asparagine synthetase deficiency","volume":"531","year":"2022","journal-title":"Clin. Chim. Acta"},{"key":"2023110623051071000_2023.10.30.564808v1.5","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymgme.2015.08.007"},{"key":"2023110623051071000_2023.10.30.564808v1.6","doi-asserted-by":"crossref","first-page":"317","DOI":"10.1016\/j.ymgme.2017.12.433","article-title":"Characterization of a novel variant in siblings with Asparagine Synthetase Deficiency","volume":"123","year":"2018","journal-title":"Mol. Genet. Metab"},{"key":"2023110623051071000_2023.10.30.564808v1.7","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.biochem.75.103004.142520"},{"key":"2023110623051071000_2023.10.30.564808v1.8","first-page":"1","article-title":"High-resolution crystal structure of human asparagine synthetase enables analysis of inhibitor binding and selectivity","volume":"2","year":"2019","journal-title":"Commun. Biol"},{"key":"2023110623051071000_2023.10.30.564808v1.9","doi-asserted-by":"crossref","first-page":"1226","DOI":"10.1002\/jimd.12408","article-title":"In vitro functional analysis of four variants of human asparagine synthetase","volume":"44","year":"2021","journal-title":"J. Inherit. Metab. Dis"},{"key":"2023110623051071000_2023.10.30.564808v1.10","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2022.102385"},{"key":"2023110623051071000_2023.10.30.564808v1.11","first-page":"559","article-title":"Analysis of Enzyme Activity and Cellular Function for the N80S and S480F Asparagine Synthetase Variants Expressed in a Child with Asparagine Synthetase Deficiency","volume":"24","year":"2023","journal-title":"Int. J. Mol. Sci"},{"key":"2023110623051071000_2023.10.30.564808v1.12","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1002\/jmd2.12356","article-title":"Characterizing asparagine synthetase deficiency variants in lymphoblastoid cell lines","volume":"64","year":"2023","journal-title":"JIMD Rep"},{"key":"2023110623051071000_2023.10.30.564808v1.13","doi-asserted-by":"crossref","first-page":"1938","DOI":"10.3390\/nu15081938","article-title":"Metabolomic Profiling of Asparagine Deprivation in Asparagine Synthetase Deficiency Patient-Derived Cells","volume":"15","year":"2023","journal-title":"Nutrients"},{"key":"2023110623051071000_2023.10.30.564808v1.14","doi-asserted-by":"publisher","DOI":"10.1093\/jn\/136.1.218S"},{"key":"2023110623051071000_2023.10.30.564808v1.15","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-020-2308-7"},{"key":"2023110623051071000_2023.10.30.564808v1.16","doi-asserted-by":"publisher","DOI":"10.1038\/nprot.2014.153"},{"key":"2023110623051071000_2023.10.30.564808v1.17","doi-asserted-by":"publisher","DOI":"10.1038\/nrg3046"},{"key":"2023110623051071000_2023.10.30.564808v1.18","doi-asserted-by":"publisher","DOI":"10.1101\/gr.176601"},{"key":"2023110623051071000_2023.10.30.564808v1.19","doi-asserted-by":"publisher","DOI":"10.1101\/gr.3577405"},{"key":"2023110623051071000_2023.10.30.564808v1.20","doi-asserted-by":"publisher","DOI":"10.1101\/gr.097857.109"},{"key":"2023110623051071000_2023.10.30.564808v1.21","doi-asserted-by":"publisher","DOI":"10.1038\/nbt.3769"},{"key":"2023110623051071000_2023.10.30.564808v1.22","doi-asserted-by":"publisher","DOI":"10.1002\/0471142905.hg0720s76"},{"key":"2023110623051071000_2023.10.30.564808v1.23","doi-asserted-by":"publisher","DOI":"10.1038\/ng.2892"},{"key":"2023110623051071000_2023.10.30.564808v1.24","doi-asserted-by":"publisher","DOI":"10.1038\/ng.3810"},{"key":"2023110623051071000_2023.10.30.564808v1.25","doi-asserted-by":"publisher","DOI":"10.1016\/j.ajhg.2016.08.016"},{"key":"2023110623051071000_2023.10.30.564808v1.26","doi-asserted-by":"crossref","first-page":"eadg7492","DOI":"10.1126\/science.adg7492","article-title":"Accurate proteome-wide missense variant effect prediction with AlphaMissense","volume":"381","year":"2023","journal-title":"Science"},{"key":"2023110623051071000_2023.10.30.564808v1.27","doi-asserted-by":"publisher","DOI":"10.1038\/S41586-021-03819-2"},{"key":"2023110623051071000_2023.10.30.564808v1.28","doi-asserted-by":"publisher","DOI":"10.1101\/2023.05.16.541009"},{"key":"2023110623051071000_2023.10.30.564808v1.29","doi-asserted-by":"publisher","DOI":"10.12688\/f1000research.29032.2"},{"key":"2023110623051071000_2023.10.30.564808v1.30","doi-asserted-by":"crossref","first-page":"687","DOI":"10.1007\/s11011-014-9618-0","article-title":"Asparagine synthetase deficiency detected by whole exome sequencing causes congenital microcephaly, epileptic encephalopathy and psychomotor delay","volume":"30","year":"2015","journal-title":"Metab. Brain Dis"},{"key":"2023110623051071000_2023.10.30.564808v1.31","doi-asserted-by":"crossref","first-page":"2071","DOI":"10.1038\/s41436-020-0919-x","article-title":"Clinical, molecular, and biochemical delineation of asparagine synthetase deficiency in Saudi cohort","volume":"22","year":"2020","journal-title":"Genet. Med"},{"key":"2023110623051071000_2023.10.30.564808v1.32","doi-asserted-by":"publisher","DOI":"10.1007\/8904_2014_405"},{"key":"2023110623051071000_2023.10.30.564808v1.33","doi-asserted-by":"crossref","first-page":"98","DOI":"10.1016\/j.pediatrneurol.2016.01.024","article-title":"Worsening of Seizures After Asparagine Supplementation in a Child with Asparagine Synthetase Deficiency","volume":"58","year":"2016","journal-title":"Pediatr. Neurol"},{"key":"2023110623051071000_2023.10.30.564808v1.34","doi-asserted-by":"crossref","first-page":"399","DOI":"10.1055\/s-0036-1586222","article-title":"Epileptic Phenotype of Two Siblings with Asparagine Synthesis Deficiency Mimics Neonatal Pyridoxine-Dependent Epilepsy","volume":"47","year":"2016","journal-title":"Neuropediatrics"},{"key":"2023110623051071000_2023.10.30.564808v1.35","doi-asserted-by":"crossref","first-page":"105","DOI":"10.1186\/s12883-016-0633-0","article-title":"Hyperekplexia, microcephaly and simplified gyral pattern caused by novel ASNS mutations, case report","volume":"16","year":"2016","journal-title":"BMC Neurol"},{"key":"2023110623051071000_2023.10.30.564808v1.36","doi-asserted-by":"crossref","first-page":"236","DOI":"10.1016\/j.braindev.2016.09.010","article-title":"The first report of Japanese patients with asparagine synthetase deficiency","volume":"39","year":"2017","journal-title":"Brain Dev"},{"key":"2023110623051071000_2023.10.30.564808v1.37","doi-asserted-by":"crossref","first-page":"200","DOI":"10.1002\/ccr3.1284","article-title":"Clinical whole exome sequencing from dried blood spot identifies novel genetic defect underlying asparagine synthetase deficiency","volume":"6","year":"2018","journal-title":"Clin. Case Rep"},{"key":"2023110623051071000_2023.10.30.564808v1.38","doi-asserted-by":"publisher","DOI":"10.1007\/8904_2016_3"},{"key":"2023110623051071000_2023.10.30.564808v1.39","doi-asserted-by":"crossref","first-page":"1889","DOI":"10.1007\/s11011-017-0073-6","article-title":"Asparagine Synthetase deficiency-report of a novel mutation and review of literature","volume":"32","year":"2017","journal-title":"Metab. Brain Dis"},{"key":"2023110623051071000_2023.10.30.564808v1.40","doi-asserted-by":"crossref","unstructured":"Schleinitz, D. et al. Novel Mutations in the Asparagine Synthetase Gene (ASNS) Associated With Microcephaly. Front. Genet. 9, (2018).","DOI":"10.3389\/fgene.2018.00245"},{"key":"2023110623051071000_2023.10.30.564808v1.41","doi-asserted-by":"crossref","first-page":"181","DOI":"10.1111\/cga.12275","article-title":"Report of four novel variants in ASNS causing asparagine synthetase deficiency and review of literature","volume":"58","year":"2018","journal-title":"Congenit. Anom"},{"key":"2023110623051071000_2023.10.30.564808v1.42","doi-asserted-by":"crossref","first-page":"100509","DOI":"10.1016\/j.ymgmr.2019.100509","article-title":"Asparagine synthetase deficiency: A novel case with an unusual molecular mechanism","volume":"21","year":"2019","journal-title":"Mol. Genet. Metab. Rep"},{"key":"2023110623051071000_2023.10.30.564808v1.43","doi-asserted-by":"crossref","first-page":"101583","DOI":"10.1016\/j.scr.2019.101583","article-title":"Generation of four induced pluripotent stem cell lines, GZWWTi001-A, GZWTZi001-A, GZWXYi001-A, and GZWXDi001-A, derived from peripheral blood mononuclear cells from a family with asparagine synthetase deficiency","volume":"41","year":"2019","journal-title":"Stem Cell Res"},{"key":"2023110623051071000_2023.10.30.564808v1.44","doi-asserted-by":"crossref","first-page":"e1235","DOI":"10.1002\/mgg3.1235","article-title":"A novel compound heterozygous missense mutation in ASNS broadens the spectrum of asparagine synthetase deficiency","volume":"8","year":"2020","journal-title":"Mol. Genet. Genomic Med"},{"key":"2023110623051071000_2023.10.30.564808v1.45","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.wneu.2019.02.130","article-title":"Cyst-Peritoneal Shunt for the Treatment of a Progressive Intracerebral Cyst Associated with ASNS Mutation: Case Report and Literature Review","volume":"127","year":"2019","journal-title":"World Neurosurg"},{"key":"2023110623051071000_2023.10.30.564808v1.46","first-page":"1","article-title":"Clinical outcomes of two patients with a novel pathogenic variant in ASNS: response to asparagine supplementation and review of the literature","volume":"6","year":"2019","journal-title":"Hum. Genome Var"},{"key":"2023110623051071000_2023.10.30.564808v1.47","doi-asserted-by":"crossref","first-page":"1343","DOI":"10.1002\/pd.5772","article-title":"Fetal MRI and ultrasound findings of a confirmed asparagine synthetase deficiency case","volume":"40","year":"2020","journal-title":"Prenat. Diagn"},{"key":"2023110623051071000_2023.10.30.564808v1.48","doi-asserted-by":"crossref","first-page":"1884","DOI":"10.1002\/humu.24101","article-title":"Rapid exome sequencing and adjunct RNA studies confirm the pathogenicity of a novel homozygous ASNS splicing variant in a critically ill neonate","volume":"41","year":"2020","journal-title":"Hum. Mutat"},{"key":"2023110623051071000_2023.10.30.564808v1.49","doi-asserted-by":"crossref","first-page":"201","DOI":"10.1055\/s-0040-1718917","article-title":"Asparagine Synthetase Deficiency with Intracranial Hemorrhage Can Mimic Molybdenum Cofactor Deficiency","volume":"52","year":"2021","journal-title":"Neuropediatrics"},{"key":"2023110623051071000_2023.10.30.564808v1.50","doi-asserted-by":"crossref","first-page":"466","DOI":"10.1684\/epd.2019.1100","article-title":"Epileptic encephalopathy with microcephaly in a patient with asparagine synthetase deficiency: a video-EEG report*","volume":"21","year":"2019","journal-title":"Epileptic. Disord"},{"key":"2023110623051071000_2023.10.30.564808v1.51","doi-asserted-by":"crossref","first-page":"107772","DOI":"10.1016\/j.clineuro.2023.107772","article-title":"Ketogenic diet-responsive drug-resistant epilepsy in a case of asparagine synthetase deficiency with a novel compound heterozygous missense variant","volume":"230","year":"2023","journal-title":"Clin. Neurol. Neurosurg"},{"key":"2023110623051071000_2023.10.30.564808v1.52","unstructured":"The PyMOL Molecular Graphics System, Version 2.5.2 Schr\u00f6dinger, LLC."}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2023.10.30.564808","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T05:27:57Z","timestamp":1768454877000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2023.10.30.564808"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,11,2]]},"references-count":52,"URL":"https:\/\/doi.org\/10.1101\/2023.10.30.564808","relation":{},"subject":[],"published":{"date-parts":[[2023,11,2]]},"subtype":"preprint"}}