Poly[(R)-3-hydroxybutyrate] (P(3HB)) single crystals, which had different morphologies with and without chain-folding surfaces, were grown from dilute solutions of chloroform and ethanol. Two types of extracellular PHB depolymerases purified from Alcaligenes faecalis T1 and Comamonas acidovorans YM1609, defined as types I and II according to the position of the lipase box in the catalytic domain, were used in the enzymatic degradation and adsorption experiments. The enzymatic degradation of P(3HB) single crystals was investigated by means of transmission electron microscopy, atomic force microscopy, and gel-permeation chromatography. Adsorption of PHB depolymerase on P(3HB) single crystals was examined using an immune-gold labeling technique. Enzymatic degradation of single crystals progressed from the edges and ends of crystals to yield the narrow cracks and the small crystal fragments along their long axis, independent of both surface morphologies of single crystals and types of extracellular PHB depolymerases. Lamellar thicknesses of single crystals and molecular weights of P(3HB) chains remained unchanged during the enzymatic hydrolysis. Adsorption of extracellular PHB depolymerase demonstrated a homogeneous distribution of enzyme molecules on the chain-folding surface of crystals. The above results suggest that the adsorption of enzyme increases with the mobility of P(3HB) chains of single crystals as a whole and that the endo-exo type attack by the active-site of PHB depolymerase takes place preferentially at the disordered chain-packing regions of crystal edges and ends rather than the chain-folding surfaces of single crystals, in spite of the difference in the structure of enzymes.