Novel Genes Encoding Hexadecanoic Acid Δ6-Desaturase Activity in a Rhodococcus sp.

cis-6-Hexadecenoic acid, a major component of human sebaceous lipids, is involved in the defense mechanism against Staphylococcus aureus infection in healthy skin and closely related to atopic dermatitis. Previously, Koike et al. (Biosci Biotechnol Biochem 64:1064-1066, 2000) reported that a mutant strain of Rhodococcus sp. produced cis-6-hexadecenoate derivatives from palmitate alkyl esters. From the mutant Rhodococcus strain, we identified and sequenced two open reading frames present in an amplified 5.7-kb region; these open reading frames encoded tandemly repeated Δ6-desaturase-like genes, Rdes1 and Rdes2. A phylogenetic tree indicated that Rdes1 and Rdes2 were different from previously known Δ6-desaturase genes, and that they formed a new cluster. Rdes1 and Rdes2 were each introduced into vectors and then expressed separately in Escherichia coli, and the fatty acid composition of the transformed cells was analyzed by gas chromatography and mass spectrometry. The amount of cis-6-hexadecenoic acid was significantly higher in Rdes1- or Rdes2-transformed E. coli cells (twofold and threefold, respectively) than in vector-only control cells. These results showed that cis-6-hexadecenoic acid was produced in E. coli cells by the rhodococcal Δ6-desaturase-like proteins.

C(35)-terpenes from Bacillus subtilis KSM 6-10.

Four C(35)-terpenes (1-4) were isolated from a liquid culture of Bacillus subtilis KSM 6-10. Compounds 1 and 2, tetraprenyl-beta-curcumene and tetraprenyl-alpha-curcumene, respectively, were previously isolated from a spore preparation of the same species, whereas 3 and 4 are new C(35)-terpenols. The C(35)-terpenols (3 and 4) possess polycyclic skeletons. We propose that the C(35)-terpenols (3 and 4) are formed by cyclization of the acyclic C(35)-terpenes (1 and 2). Because only trace amounts of C(35)-terpenols 3 and 4 were found in the spore in contrast to the vegetative cells of B. subtilis KSM 6-10, it is assumed that the expression of terpene cyclase is not related to sporulation, as has been observed by other researchers in a different strain.

Hyperproduction of sebaceous cis-6-hexadecenoic acid by esterase-reduced mutant of Rhodococcus sp. strain.

cis-6-Hexadecenoic acid is a major component of human sebaceous lipids that is involved in skin self-sterilization and atopic dermatitis amelioration. It can be prepared by hydrolysis of isopropyl cis-6-hexadecenoate produced by resting cells of Rhodococcus sp. strain KSM-MT66. To devise an economical industrial-scale process for the production of this rare fatty acid, we optimized the conditions for growing rhodococcal cells. Mg(2+) and Fe(2+) ions are essential for the efficient production of isopropyl cis-6-hexadecenoate. To further increase the production of isopropyl cis-6-hexadecenoate, we created a mutant strain (T64) with reduced esterase activity by random mutagenesis using UV irradiation of MT66. Under an optimized condition, the mutant T64 produced more than 60 g l(-1) isopropyl cis-6-hexadecenoate in a 4-d cultivation, corresponding to about 52 g l(-1)cis-6-hexadecenoate.

Deficient production of hexadecenoic acid in the skin is associated in part with the vulnerability of atopic dermatitis patients to colonization by Staphylococcus aureus.

BACKGROUND AND OBJECTIVES: As one of the major skin fatty acids, cis-6-hexadecenoic acid (C16:1Delta6) exhibits a specific antibacterial activity and might play a specific role in the defense mechanism against Staphylococcus aureus, in healthy subjects whereas S. aureus frequently colonizes the skin of patients with atopic dermatitis (AD). METHODS: Fatty acid composition of sebum at the recovery level was analyzed by gas chromatography and S. aureus colonizing the skin was assessed by the 'cup-scrub' method (9 patients and 10 healthy controls). To evaluate in vivo effect of C16:1Delta6 on colonization, C16:1Delta6 was applied for 2 weeks on the upper arm skin of another group of AD patients (11 patients). RESULTS: Analysis of sebum lipids revealed that there is a significant lower free C16:1Delta6 content in nonlesional skin from AD patients compared with healthy controls. This lower content is also accompanied by a significantly lower level of C16:1Delta6 in the total fatty acid composition of sebum (analyzed following hydrolysis). The lower level of free C16:1Delta6 correlated significantly (R(2) = 0.41, p < 0.01) with the numbers of S. aureus colonizing nonlesional skin. Topical application of free C16:1Delta6 on the skin of AD patients for 2 weeks abolished the markedly increased bacterial count in 6 out of the 8 AD patients tested. CONCLUSIONS: Free C16:1Delta6 may be involved in the defense mechanism against S. aureus in healthy skin and this deficit triggers the susceptibility of the skin to colonization by S. aureus in AD.