Gastric digestion of proteins is influenced by multiple factors including microstructure, mechanical properties and structure breakdown during mastication. The interplay between these factors affects protein digestion but is underexplored. This study aimed to investigate the contribution of microstructure, mechanical properties and macrostructure breakdown on in vitro whey protein gastric digestion. Whey protein isolate (WPI) was mixed with different types of polysaccharides (κ-carrageenan, ι-carrageenan, pectin) at various concentrations to obtain heat- or acid-induced gels with distinct microstructures (homogeneous, coarse stranded, protein continuous and bi-continuous) and Young's moduli (E, 19–165 kPa). Structural breakdown during mastication was mimicked crudely by cutting single gel cylinders into several smaller cubes to increase the total surface area by a factor of 2.65. In vitro gastric digestion was measured using the INFOGEST 2.0 protocol with minor modifications. Homogeneous heat-induced WPI/κ-carrageenan gels showed the highest digestion rate followed by protein continuous, coarse stranded and bi-continuous heat-induced WPI/κ-carrageenan gels with similar E. A 1.47-fold increase in E decreased the digestion rate of acid-induced homogeneous WPI/ι-carrageenan gels by a factor of 0.22. In contrast, a 1.13–1.83-fold increase in E barely changed the digestion rate of acid-induced protein continuous WPI/pectin gels. A 2.65-fold increase in total surface area increased the digestion rate of all gels by a factor of 1.35–2.54 depending on microstructure and mechanical properties. We conclude that the microstructure of protein gels affects in vitro protein gastric digestion and the impact of Young's modulus on in vitro protein gastric digestion depends strongly on the microstructure of protein gels.