The inhibitory effect of toxic byproducts mainly derived from the pretreatment of lignocellulosic biomass is one of the limiting factors hindering industrial application. In this work, a simple yet practicable aeration strategy was proposed to enhance microbial tolerance to inhibitors, including formic acid, acetic acid, furfural, and 5-hydroxymethyl furfural (5-HMF). Batch fermentation by Kluyveromyces marxianus CICC 1727-5 using xylose and glucose alone and in combination under different aeration conditions coupled with multiple inhibitors was performed. The fermentation time and residual xylose significantly decreased with high xylitol production (0.7 g/g) and high ethanol production (0.42 g/g) at an aeration rate of 0.5 vvm. The postglucose effect was effectively relieved during co-fermentation by the microaeration strategy. Transcriptomics analysis was performed to determine the mechanism of inhibitor resistance. Transcription of genes related to DNA replication, protein processing, carbon metabolism, biosynthesis of the ribosome, amino acids, peroxidase, sterols, and unsaturated fatty acids were significantly upregulated accompanied by downregulated expression of genes related to autophagy under aeration conditions in the presence of multiple inhibitors. These results provide a valuable reference for future studies on biofuel fermentation from lignocellulosic biomass by a facultative fermenting microbe.