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Updated: Jan 14, 2026
A Novel Saturation Mutagenesis Approach: Single Step Characterization of Regulatory Protein Binding Sites in RNA Using Phosphorothioates
Published on: August 21, 2018
Characterization and expression of human bifunctional 3'-phosphoadenosine 5'-phosphosulphate synthase isoforms
Hirotoshi Fuda1, Chikara Shimizu, Young C Lee
1Section on Steroid Regulation, Endocrinology and Reproduction Research Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892-4510, USA.
PAPS synthase 2 subtypes exhibit significantly higher activity than PAPS synthase 1, crucial for sulfonation. This difference helps explain human growth disorders linked to PAPS synthase 2 mutations, particularly in developing cartilage.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Sulfonation is vital for growth and development, relying on 3'-phosphoadenosine 5'-phosphosulphate (PAPS) as the sulfonate donor.
- PAPS is synthesized by PAPS synthase (PS) from ATP and inorganic sulfate.
- Humans have two PAPS synthase genes, with PAPS synthase 2 (PS2) having alternative splicing, creating subtypes.
Purpose of the Study:
- To investigate if amino acid differences between human PAPS synthase isoenzymes and subtypes lead to functional variations.
- To explore the catalytic efficiency and expression patterns of PAPS synthase 1 (PS1) and PS2 subtypes.
- To resolve the apparent contradiction between ubiquitous PS1 expression and growth disorders caused by PS2 mutations.
Main Methods:
- Enzyme activity assays were performed to compare the specific activity and catalytic efficiency (kcat/Km) of PS1 and PS2 subtypes.
- Gene expression analysis was conducted in various human tissues and in guinea-pig cartilage at different developmental stages.
- Comparative analysis of enzyme kinetics and tissue-specific expression patterns.
Main Results:
- PAPS synthase 2 subtypes demonstrated 10- to 15-fold higher specific activity compared to PAPS synthase 1.
- PS2 subtypes exhibited 3- to 6-fold greater catalytic efficiency (kcat/Km) for both ATP and inorganic sulfate substrates than PS1.
- While PS1 is ubiquitously expressed, PS2 expression is tissue-specific. Notably, immature guinea-pig cartilage, unlike mature cartilage, shows high PS2 and low PS1 expression.
Conclusions:
- Significant catalytic differences exist between human PAPS synthase isoenzymes, with PS2 subtypes being more efficient.
- The tissue-specific and developmentally regulated expression of PS2, particularly in cartilage, is critical for normal growth.
- The findings resolve the enigma surrounding PS2 mutations causing dwarfing disorders, highlighting the importance of PS2 in skeletal development.
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