The c.521T>A variant in PAH is a missense variant predicted to cause substitution of Isoleucine by Asparagine at amino acid 174 (p.Ile174Asn). At least one patient with this variant displayed plasma phenylalanine > 1.2 mmol/L, which is highly specific for PAH deficiency; BH4 deficiency was excluded by analysis of dihydropteridine reductase activity in red blood cells, biopterin loading test and/or pteridine analysis in urine (PP4_Moderate; PMID: 21307867, 23842451). Additional individuals with PAH deficiency were compound heterozygous for the variant and a pathogenic variant (phase unknown; c.1315+1G>A, p.P211T, 1.0 point, PMID: 23842451. The highest population minor allele frequency in gnomAD v4.1 is 0.00008010, which is lower than the ClinGen PAH threshold (<0.0002) for PM2_Supporting, meeting this criterion (PM2_Supporting). The computational predictor REVEL gives a SCORE of 0.929, which is between the threshold of 0.773 - 0.932, evidence that correlates with moderate impact to PAH function (PP3_moderate). In summary, this variant meets criteria to be classified as likely pathogenic for PAH deficiency based on the ACMG/AMP criteria applied, as specified by the ClinGen PAH VCEP: PP4_Moderate, PM2_supporting, PM3, PP3_moderate.