<jats:title>Abstract</jats:title><jats:p>In recent years, biotechnological conversion of the alternative carbon source acetate has attracted much attention. So far, acetate has been mainly used for microbial production of bioproducts with bulk applications. In this study, we aimed to investigate the potential of acetate as carbon source for heterologous protein production using the acetate‐utilizing platform organism <jats:italic>Corynebacterium glutamicum</jats:italic>. For this purpose, expression of model protein eYFP with the promoter systems T7<jats:italic>lac</jats:italic> and <jats:italic>tac</jats:italic> was characterized during growth of <jats:italic>C. glutamicum</jats:italic> on acetate as sole carbon source. The results indicated a 3.3‐fold higher fluorescence level for acetate‐based eYFP production with T7 expression strain MB001(DE3) pMKEx2‐<jats:italic>eyfp</jats:italic> compared to MB001 pEKEx2‐<jats:italic>eyfp</jats:italic>. Interestingly, comparative <jats:italic>eyfp</jats:italic> expression studies on acetate or glucose revealed an up to 83% higher biomass‐specific production for T7 RNAP‐dependent eYFP production using acetate as carbon source. Furthermore, high‐level protein accumulation on acetate was demonstrated for the first time in a high cell density cultivation process with pH‐coupled online feeding control, resulting in a final protein titer of 2.7 g/L and product yield of 4 g per 100 g cell dry weight. This study presents a first proof of concept for efficient microbial upgrading of potentially low‐cost acetate into high‐value bioproducts, such as recombinant proteins.</jats:p>