BackgroundIn the sporadic form of amyotrophic lateral sclerosis (ALS), the pathogenicity of rare variants in the causative genes characterizing the familial form remains largely unknown. To predict the pathogenicity of such variants, in silico analysis is commonly used. In some ALS causative genes, the pathogenic variants are concentrated in specific regions, and the resulting alterations in protein structure are thought to significantly affect pathogenicity. However, existing methods have not taken this issue into account. To address this, we have developed a technique termed MOVA (method for evaluating the pathogenicity of missense variants using AlphaFold2), which applies positional information for structural variants predicted by AlphaFold2. Here we examined the utility of MOVA for analysis of several causative genes of ALS.MethodsWe analyzed variants of 12 ALS-related genes (TARDBP,FUS,SETX,TBK1,OPTN,SOD1,VCP,SQSTM1,ANG,UBQLN2,DCTN1, andCCNF) and classified them as pathogenic or neutral. For each gene, the features of the variants, consisting of their positions in the 3D structure predicted by AlphaFold2, pLDDT score, and BLOSUM62 were trained into a random forest and evaluated by the stratified fivefold cross validation method. We compared how accurately MOVA predicted mutant pathogenicity with other in silico prediction methods and evaluated the prediction accuracy atTARDBPandFUShotspots. We also examined which of the MOVA features had the greatest impact on pathogenicity discrimination.ResultsMOVA yielded useful results (AUC ≥ 0.70) forTARDBP,FUS,SOD1,VCP, andUBQLN2of 12 ALS causative genes. In addition, when comparing the prediction accuracy with other in silico prediction methods, MOVA obtained the best results among those compared forTARDBP,VCP,UBQLN2, andCCNF. MOVA demonstrated superior predictive accuracy for the pathogenicity of mutations at hotspots ofTARDBPandFUS. Moreover, higher accuracy was achieved by combining MOVA with REVEL or CADD. Among the features of MOVA, the x, y, and z coordinates performed the best and were highly correlated with MOVA.ConclusionsMOVA is useful for predicting the virulence of rare variants in which they are concentrated at specific structural sites, and for use in combination with other prediction methods.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12859-023-05338-5.