Glycogen-free synthase I from human polymorphonuclear leukocytes is activated by its own substrate, glycogen, in a slow, time-dependent process (hysteretic activation). This lag in response to addition of glycogen depends on the concentration of glycogen, pH and temperature. At pH 7.4 and at a temperature of 30 degrees C, the half-time of activation t 1/2 decreases from 89 min at 0.004 mg/ml glycogen to 6 min at 25 mg/ml. The activation is accelerated by increasing temperature and pH, but is not influenced by enzyme concentration, glucose 6-phosphate, UDP, high ionic strength, EDTA, mercaptoethanol, glucose, sucrose or amylase limit dextrin. The Km for UDP-glucose (0.024 mM) and the activity ratio were unchanged during the activation process. The activation can be described by vt = vf + (vo - vf) e-kt where vt, vf and vo are velocities at times t, O and infinity and k is a complex rate constant. Evidence from ultracentrifugation and kinetic studies is presented to substantiate the hypothesis that the underlying mechanism is a simple biolecular process: enzyme + glycogen in equilibrium enzyme-glycogen complex, with the dissociation constant Ks = 0.003 mg/ml. The hysteretic activation may become rate-limiting during experiments in vitro with synthase. The possibility of a physiological role in glycogen metabolism, perhaps in the form of a concerted hysteresis with H+ is discussed.