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PreprintCited by 2Biochemical and Molecular ResearchEnzyme Structure and FunctionProtein Structure and Dynamics

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Disruption of CAD oligomerization by pathogenic variants

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Del Caño-Ochoa, Francisco, Ramadane-Morchadi, Lobna, Eixerés, Lluís, Moreno-Morcillo, María, Fernández-Leiro, Rafael, Ramón-Maiques, Santiago·2024·Journal of Molecular Biology
DOI 10.1101/2024.07.30.605799

Preprint — published as 10.1016/j.jmb.2024.168832 · Paper page

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Abstract

CAD is a multi-enzymatic protein essential for initiating the de novo biosynthesis of pyrimidine nucleotides, forming large hexamers whose structure and function are not fully understood. Defects in CAD result in a severe neurometabolic disorder that is challenging to diagnose. We developed a cellular functional assay to identify defective CAD variants, and in this study, we characterized five pathogenic missense mutations in CAD’s dihydroorotase (DHO) and aspartate transcarbamylase (ATC) domains. All mutations impaired enzymatic activities, with two notably disrupting the formation of DHO dimers and ATC trimers. Combining crystal structures and AlphaFold predictions, we modeled the hexameric CAD complex, highlighting the central role of the DHO and ATC domains in its assembly. Our findings provide insight into CAD’s stability, function, and organization, revealing that correct oligomerization of CAD into a supramolecular complex is required for its function in nucleotide synthesis and that mutations affecting this assembly are potentially pathogenic.

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Metadata

Authors
Del Caño-Ochoa, F., Ramadane-Morchadi, L., Eixerés, L., Moreno-Morcillo, M., Fernández-Leiro, R., & Ramón-Maiques, S.
Affiliations
  • Consejo Superior de Investigaciones Científicas (ES)
  • Spanish National Cancer Research Centre (ES)
ORCID
Venue
Journal of Molecular Biology
Publisher
openRxiv
Published
2024-07-30
Volume
436
Issue
23
Pages
168832-168832
ISSN
0022-2836 (ISSN-L)
DOI
10.1101/2024.07.30.605799
OpenAlex
W4403628001
Cited by
2
References
1
Citations by year
2025: 2
FWCI
0.27
Type
journal article
Language
en
Open access
Licence
Locations
  • Journal of Molecular Biology (publishedVersion, cc-by-nc-nd)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
  • DIGITAL.CSIC (Spanish National Research Council (CSIC)) (acceptedVersion)
Funders
  • Fundación Ramón Areces
  • Ministerio de Ciencia, Innovación y Universidades
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Molecular Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Enzyme Structure and Function
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
Keywords
protein trimerization, protein dimerization, pyrimidine synthesis, neurometabolic disorders
MeSH terms
Protein Domains, Aspartate Carbamoyltransferase, Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing), Dihydroorotase, Humans, Models, Molecular, Pentosyltransferases, Protein Conformation, Crystallography, X-Ray, Mutation, Missense, Protein Multimerization

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