Nanolayer TiAlN/TiSiN coating is one of the most advanced contemporary protective coatings. It has been applied for protection of machining tools, forming tools, and die casting tools. However, due to its versatile properties, there is a high potential for broadening its application; for example, for protection of biomedical implants. Each application requires specific base materials, for example cold working steels are used for forming, while stainless steels are applied for biomedical purposes. Different materials and their pre-treatment might result in different coating properties even if coating was conducted in a single batch. Real tools and components have complex geometries, and as such require a multiple-axis rotation during the deposition. Among other properties, grain morphology and surface topography are of great importance in a real application. Since systematic studies on the effect of substrate materials and rotation during deposition on these properties are very scarce, in this article we studied TiAlN/TiSiN coating magnetron sputtered on five different substrates, prepared with 1-, 2-, and 3-fold rotations. Cold-work tool steel (X153CrMoV12), hot-work tool steel (X37CrMoV5-1), plasma-nitrided hot-work tool steel, surgical stainless steel (X2CrNiMo18-15-3), and cemented carbide (WC/Co) were used as substrate materials. Three-dimensional stylus profilometry and atomic force microscopy were used for evaluation of micro and nano topography. The coated surgical steel has the highest roughness (Sa) which corresponds to the highest number of coating growth defects. However, the size of the individual growth defects was considerably smaller for this substrate than for other substrate materials. The observed difference is linked to differences in the concentration of specific carbides contained in a specific steel. Since different carbides have different polishing and ion-etching rates, coatings on different steels may have different concertation of defects. Columnar grain analysis revealed that coating on surgical steel exhibited the smallest column diameter (125 nm) and their highest uniformity. Column diameter on other substrates is around 215 nm, while hot-working tool steel exhibited the largest columns (235 nm). Such findings suggest that the same coating may exhibit different mechanical properties on different substrates. Coatings produced with the higher degree of rotation (2-fold, 3-fold) have fewer defects and a smoother surface. There was no clear trend between columnar grain size and the number of rotational degrees.