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Hydroxypropyl-β-cyclodextrin (HP-β-CD) enhances steroid 1-dehydrogenation biotransformation by Arthrobacter simplex. In this work, HP-β-CD-induced improvement of A. simplex CPCC 140451 cell envelope permeability which had positive effects on the steroid bioconversion was confirmed by a comparative investigation which showed a lower dehydrogenase activity and higher cell permeability of the cells after being incubated with HP-β-CD. Atomic force microscopy and transmission electron microscopy micrographs showed that HP-β-CD altered the size, sharpness, and surface structure of the cell envelope. The analysis of lipid composition revealed that the proportion of extractable lipids decreased and the fatty acids profile was considerably altered. The contents of unsaturated fatty acids and long-chain fatty acids were reduced by 11.77 and 14.98 %, respectively. The total leakage of protein level increased to 8 %. Proteins belonging to the ATP-binding cassette superfamily and major facilitator superfamily were observed outside the cell. These alterations can explain the change of permeability on the molecular level under HP-β-CD treatment. Results showed the material basis and mechanisms underlying the cellular changes, thus most likely contributing to the conversion rate in addition to cyclodextrins known effects on substrate solubility. [ABSTRACT FROM AUTHOR]