Microbial marine exopolysaccharides (EPS) are one of the essential and functional commercial-grade products and have several biotechnological applications. In this study, marine Pseudoalteromonas shioyasakiensis SE strain isolated from Alexandria, Egypt, was identified and characterized biochemically and molecularly. A novel exopolysaccharide (EPSSE) was extracted and isolated. The molecular weight (Mw) of EPSSE, as determined by structural characterization, is a glycosidic heteropolysaccharide with 8 kDa and has no uronic acid. Additionally, the results showed that EPSSE was a basic EPS composed of sucrose, glucose, and fructose with the ratio of ≈ 5:18:1 respectively. The maximum antioxidant activity was 12.43% at 2000 µg/mL. The anti-inflammatory activity of EPSSE indicated that it has significant inhibitory activity toward cyclooxygenase (COX-1) and (COX-2) activities with IC50 values of 19.24 ± 1.14 µg/mL and 4.12 ± 0.21) µg/mL, respectively. Furthermore, EPSSE exhibited considerable inhibitory activity toward the proliferation of WI-38, MCF-7 cells, HepG-2 cells and A-549 cell lines with IC50 values of 756.2 ± 36.8 µg/mL, 480.2 ± 21, 244.1 ± 9.3 µg/mL and 398.3 ± 20.9, respectively. This study revealed that EPS produced by the marine P. shioyasakiensis SE strain could be considered an important source of pharmacologically active compounds that would be useful for the healthcare system.