Chemically inert hydrocarbons are the primary feedstocks used in the petrochemical industry and can be converted into more intricate and valuable chemicals. However, two major challenges impede this conversion process: selective activation of C−H bonds in hydrocarbons and systematic functionalization required to synthesize complex structures. To address these issues, we developed a multi-enzyme cascade conversion system based on internal cofactor and H₂O₂ recycling to achieve the one-pot deep conversion from heptane to chiral (S)-2-aminoheptanoic acid under mild conditions. First, a hydrogen-borrowing-cycle-based NADH regeneration method and H₂O₂ in situ generation and consumption strategy were applied to realize selective C−H bond oxyfunctionalization, converting heptane into 2-hydroxyheptanoic acid. Integrating subsequent reductive amination driven by the second hydrogen-borrowing cycle, (S)-2-aminoheptanoic acid was finally accumulated at 4.57 mM with ee_p>99 %. Hexane, octane, 2-methylheptane, and butylbenzene were also successfully converted into the corresponding chiral amino acids with ee_p>99 %. Overall, the conversion system employed internal cofactor and H₂O₂ recycling, with O₂ as the oxidant and ammonium as the amination reagent to fulfill the enzymatic conversion from chemically inert hydrocarbons into chiral amino acids under environmentally friendly conditions, which is a highly challenging transformation in traditional organic synthesis.