This study investigates the biochemical properties of two xylanases, ZgXyn10A and CaXyn10B, which are members of the glycoside hydrolase family 10 (GH10) and originate from the marine Bacteroidetes species and , respectively. Utilizing an auto-induction expression system in , high-purity recombinant forms of these enzymes were successfully produced. Biochemical assays revealed that ZgXyn10A and CaXyn10B exhibit optimal activities at 40 °C and 30 °C, respectively, and demonstrate a high sensitivity to temperature fluctuations. Unlike conventional low-temperature enzymes, these xylanases retain only a fraction of their maximal activity at lower temperatures. To gain deeper insights into the structural and functional properties of these marine xylanases, two thermostable GH10 xylanases, TmxB and CoXyn10A, which share comparable amino acid sequence identity with ZgXyn10A and CaXyn10B, were selected for structural comparison. All four marine xylanases share a nearly similar three-dimensional structural topology. Molecular dynamics simulation indicated a striking difference in structural fluctuations between the low-temperature and thermostable xylanases, as evidenced by the distinct root mean square deviation values. Moreover, root mean square fluctuation analysis specifically identified the β3-α3 and β7-α7 loop regions within the substrate-binding cleft as crucial determinants of the temperature characteristics of these GH10 xylanases. Our findings establish loop dynamics as a key evolutionary driver in the thermal adaptation of GH10 xylanases and propose a loop engineering strategy for the development of industrial biocatalysts with tailored temperature responses, particularly for lignocellulosic biomass processing under moderate thermal conditions.