Analysis of the Qatari R336C cystathionine β-synthase protein in mice

Area of Science:

• Biochemistry
• Genetics
• Metabolic Disorders

Background:

• Classical homocystinuria is an inherited metabolic disorder resulting from mutations in the cystathionine beta-synthase (CBS) gene.
• A specific founder mutation, c.1006C>T (p.R336C), is prevalent in Qatar, causing a severe, pyridoxine-unresponsive form of the disease.

Purpose of the Study:

• To create and characterize a mouse model expressing the severe human p.R336C CBS mutation.
• To investigate the molecular mechanisms underlying the p.R336C mutation's effect on CBS protein stability and enzyme activity.

Main Methods:

• Generation of a mouse model (Tg-R336C Cbs ^-/-) with a null Cbs gene and a zinc-inducible human CBS transgene.
• Analysis of serum and liver homocysteine levels, CBS protein levels, and enzyme activity in treated and untreated mice.
• In vitro enzymatic studies to assess substrate and cofactor binding affinities of the p.R336C CBS variant.

Main Results:

• Zinc-induced expression of p.R336C CBS in mice led to severe hyperhomocysteinemia.
• The p.R336C mutation significantly reduced CBS protein stability and enzyme activity in mouse liver.
• Proteasome inhibitor treatment partially restored CBS activity but did not fully correct serum homocysteine levels.
• In vitro studies revealed that p.R336C CBS has reduced serine binding affinity and increased pyridoxal phosphate dependence.

Conclusions:

• The p.R336C mutation impairs CBS protein stability and alters substrate/cofactor binding, contributing to severe classical homocystinuria.
• This mouse model provides a valuable tool for studying the pathophysiology of this specific CBS deficiency and for testing therapeutic strategies.