The structural origin of metabolic quantitative diversity

Metabolomics is emerging as an indispensable method for investigating the causes of diseases, because metabolite levels in biofluids are significantly influenced by various genetic and environmental factors. Recent technological advances have made it possible to implement the genome wide association study (GWAS) of metabolic traits to investigate genetic effects on blood metabolite levels1,2,3,4,5,6. Many blood metabolites appear to be associated with genetic loci, suggesting that normal blood metabolite levels may be influenced by genetic polymorphisms. In most of previous GWAS, individual samples were genotyped exploiting commercial array systems that contain a limited number of single nucleotide polymorphisms (SNPs), resulting in frequent difficulties in identifying the causal polymorphisms influencing the metabolite levels, leaving the question open as to how these polymorphisms affect the blood metabolite levels at a molecular or catalytic level. As elucidation of the relationship between quantitative diversity of human blood metabolites and structural diversity of enzymes caused by variants in human population is critically important to understand the mechanisms how human metabolic individuality is defined, in this study we performed a large-scale cohort association study of metabolomics-genomics to investigate the relationship between structural variations in enzymes and metabolic phenotypes in human. We analyzed the metabolites of plasma collected from 512 participants in a population cohort conducted by Tohoku Medical Megabank organization (ToMMo) by nuclear magnetic resonance (NMR) spectroscopy. We obtained the metabolite profiling of plasma and analyzed the correlation among the quantified metabolites. We also performed an association study of plasma metabolites using whole-genome sequence dataset from all participants7 to elucidate true causal non-synonymous variants, instead of using SNP array data. We identified five metabolites associated with non-synonymous variants in five metabolic enzymes, four of which were previously reported to be involved in metabolic diseases. To clarify the relationship between the variants and functional activity of related enzymes, we performed structural analysis of the five non-synonymous variants and found that they are not located in catalytic center regions, but located in peripheral regions or in regulatory domains, indicating that these variants retain only moderate impact on their corresponding enzymatic activities. Therefore, we further analyzed variants in these enzymes and found two individuals with larger changes of metabolite levels. These individuals have much more rare variants of one enzyme gene that cause non-synonymous variants located in closer proximity to the catalytic site, indicating that they cause larger functional impacts than the moderate variant. Whereas many studies have been conducted to clarify the relationship between gene variants and functional activities of proteins, our results unequivocally demonstrate that variant frequency, structural location, and effect for phenotype correlate with each other in human population. We expect that our approach is versatile to discover further associations of metabolites with diseases, as even a moderate variant can be detected as being associated with a significant change of plasma metabolite levels.