Isolation of Animal Cell Mutants Defective in Long-chain Fatty Aldehyde Dehydrogenase

The kinetic mechanism of a long-chain aldehyde dehydrogenase that is induced during the development of bioluminescence in Beneckea harveyi has been investigated. Parallel lines were obtained in Lineweaver–Burk plots with NAD+ and long-chain aldehydes (heptaldehyde, nonylaldehyde). However, product and dead-end inhibitor studies, substrate protection (NAD+, aldehyde) against inactivation with N-ethylmaleimide, and in particular, a noncompetitive substrate inhibition pattern with aldehyde at high concentrations showed that aldehyde dehydrogenase had a sequential mechanism. The data were consistent with a nonrapid equilibrium random mechanism with a preferred order of addition of substrates (NAD+, aldehyde) and an ordered release of products with NADH release being the last and rate-limiting step in the reaction, a mechanism very similar to that found for short-chain mammalian aldehyde dehydrogenases. The present experiments emphasize the caution that must be taken in interpreting parallel patterns in initial velocity experiments, as well as the difficulty in classifying sequential enzyme mechanisms as either strictly ordered or random.

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Impaired skin barrier function due to reduced ω- O -acylceramide levels in a mouse model of Sjögren–Larsson syndrome

Molecular and Cellular Biology ◽

10.1128/mcb.00352-21 ◽

2021 ◽

Author(s):

Koki Nojiri ◽

Shuhei Fudetani ◽

Ayami Arai ◽

Takuya Kitamura ◽

Takayuki Sassa ◽

...

Keyword(s):

Molecular Mechanism ◽

Aldehyde Dehydrogenase ◽

Barrier Function ◽

Skin Barrier ◽

Skin Barrier Function ◽

Fatty Aldehyde ◽

Chain Base ◽

Long Chain Base ◽

Larsson Syndrome ◽

Long Chain

Sjögren–Larsson syndrome (SLS) is an inherited neurocutaneous disorder whose causative gene encodes the fatty aldehyde dehydrogenase ALDH3A2. To date, the detailed molecular mechanism of the skin pathology of SLS has remained largely unclear. We generated double knockout (DKO) mice for Aldh3a2 and its homolog Aldh3b2 (a pseudogene in humans). These mice showed hyperkeratosis and reduced fatty aldehyde dehydrogenase activity and skin barrier function. The levels of ω- O -acylceramides (acylceramides), which are specialized ceramides essential for skin barrier function, in the epidermis of DKO mice were about 60% of those in wild type mice. In the DKO mice, levels of acylceramide precursors (ω-hydroxy ceramides and triglycerides) were increased, suggesting that the final step of acylceramide production was inhibited. A decrease in acylceramide levels was also observed in human immortalized keratinocytes lacking ALDH3A2 . Differentiated keratinocytes prepared from the DKO mice exhibited impaired long-chain base metabolism. Based on these results, we propose that the long-chain-base–derived fatty aldehydes that accumulate in DKO mice and SLS patients attack and inhibit the enzyme involved in the final step of acylceramide. Our findings provide insight into the pathogenesis of the skin symptoms of SLS, i.e., decreased acylceramide production, and its molecular mechanism.

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