<p>Alginate is a polysaccharide predominantly synthesized by brown algae and bacteria. It is a linear polymer composed of β-D-mannuronic acid (M) and its C5 reverse isomer α-L-guluronic acid (G). Currently, there is a significant demand for high-G-content alginate in the market. Mannuronan C5-epimerase (MC5E) is one important enzyme in the alginate synthesis as it catalyzes epimerization of β-D-mannuronic acid (M) to α-L-guluronic acid (G) at the polyM polymer level. We screened and isolated one high expression <i>MC5E</i> sequence from the <i>Saccharina japonica</i> genome database and demonstrated that <i>SjMC5E1</i> could epimerize and increase the content of G residues in alginate. Additionally, we detected the mannuronan C5-epimerase activity of <i>SjMC5E1</i> was optimal at 37 °C in the temperature range of 15–50 °C, the <i>F</i>_G was 0.30 and the M/G ratio of alginate was the lowest at 2.32. The biochemical characterization of <i>SjMC5E1</i> provides valuable insights into its role in alginate biosynthesis, particularly in the conversion of M to G. These findings contribute to a better understanding of the enzymatic mechanisms underlying alginate synthesis and may support future efforts to optimize alginate production. However, further studies are needed to fully explore the potential of <i>SjMC5E1</i> in tailoring alginates with specific properties for industrial or biomedical applications.</p>