<jats:title>ABSTRACT</jats:title> <jats:p> <jats:named-content content-type="genus-species">Corynebacterium glutamicum</jats:named-content> was metabolically engineered to produce 4-hydroxybenzoic acid (4-HBA), a valuable aromatic compound used as a raw material for the production of liquid crystal polymers and paraben. <jats:named-content content-type="genus-species">C. glutamicum</jats:named-content> was found to have a higher tolerance to 4-HBA toxicity than previously reported hosts used for the production of genetically engineered 4-HBA. To obtain higher titers of 4-HBA, we employed a stepwise overexpression of all seven target genes in the shikimate pathway in <jats:named-content content-type="genus-species">C. glutamicum</jats:named-content> . Specifically, multiple chromosomal integrations of a mutated <jats:italic>aroG</jats:italic> gene from <jats:named-content content-type="genus-species">Escherichia coli</jats:named-content> , encoding a 3-deoxy- <jats:sc>d</jats:sc> -arabinoheptulosonic acid 7-phosphate (DAHP) synthase, and wild-type <jats:italic>aroCKB</jats:italic> from <jats:named-content content-type="genus-species">C. glutamicum</jats:named-content> , encoding chorismate synthase, shikimate kinase, and 3-dehydroquinate synthase, were effective in increasing product titers. The last step of the 4-HBA biosynthesis pathway was recreated in <jats:named-content content-type="genus-species">C. glutamicum</jats:named-content> by expressing a highly 4-HBA-resistant chorismate pyruvate-lyase (UbiC) from the intestinal bacterium <jats:named-content content-type="genus-species">Providencia rustigianii</jats:named-content> . To enhance the yield of 4-HBA, we reduced the formation of by-products, such as 1,3-dihydroxyacetone and pyruvate, by deleting <jats:italic>hdpA</jats:italic> , a gene coding for a haloacid dehalogenase superfamily phosphatase, and <jats:italic>pyk</jats:italic> , a gene coding for a pyruvate kinase, from the bacterial chromosome. The maximum concentration of 4-HBA produced by the resultant strain was 36.6 g/liter, with a yield of 41% (mol/mol) glucose after incubation for 24 h in minimal medium in an aerobic growth-arrested bioprocess using a jar fermentor. To our knowledge, this is the highest concentration of 4-HBA produced by a metabolically engineered microorganism ever reported. <jats:p> <jats:bold>IMPORTANCE</jats:bold> Since aromatic compound 4-HBA has been chemically produced from petroleum-derived phenol for a long time, eco-friendly bioproduction of 4-HBA from biomass resources is desired in order to address environmental issues. In microbial chemical production, product toxicity often causes problems, but we confirmed that wild-type <jats:named-content content-type="genus-species">C. glutamicum</jats:named-content> has high tolerance to the target 4-HBA. A growth-arrested bioprocess using this microorganism has been successfully used for the production of various compounds, such as biofuels, organic acids, and amino acids. However, no production method has been applied for aromatic compounds to date. In this study, we screened for a novel final reaction enzyme possessing characteristics superior to those in previously employed microbial 4-HBA production. We demonstrated that the use of the highly 4-HBA-resistant UbiC from the intestinal bacterium <jats:named-content content-type="genus-species">P. rustigianii</jats:named-content> is very effective in increasing 4-HBA production.