Chemical synthesis of lactones from cycloalkanes is a multi-step process challenged by limitations in reaction efficiency (conversion and yield), atom economy (by-products) and environmental performance. A heterologous pathway comprising novel enzymes with compatible kinetics was designed in Pseudomonas taiwanensis VLB120 enabling in-vivo cascade for synthesizing lactones from cycloalkanes. The respective pathway included cytochrome P450 monooxygenase (CHX), cyclohexanol dehydrogenase (CDH), and cyclohexanone monooxygenase (CHXON) from Acidovorax sp. CHX100. Resting (non-growing) cells of the recombinant host P. taiwanensis VLB120 converted cyclohexane, cyclohexanol, and cyclohexanone to E-caprolactone at 22, 80-100, and 170Ug(CDW)(-1), respectively. Cyclohexane (5mM) was completely converted with a selectivity of 65% for E-caprolactone formation in 2hr without accumulation of intermediate products. Promiscuity of the whole-cell biocatalyst gave access to analogous lactones from cyclooctane and cyclodecane. A total product concentration of 2.3gL(-1) and a total turnover number of 36,720 was achieved over 5hr with a biocatalyst concentration of 6.8g(CDW)L(-1).