The enzyme UDP-N-acetylglucosamine:dolichyl phosphate, N-acetylglucosamine-1-phosphate transferase (GlcNAc-1-P transferase), the first committed step in the dolichol-linked oligosaccharide pathway for glycoprotein biosynthesis, has been studied in developing rat brain. The enzyme was shown to be localized in microsomes, particularly heavy microsomes, and to be activated by Mg2+ and inhibited by tunicamycin. Study of the enzyme with brain development demonstrated two prominent findings. First, the accentuation of enzymatic activity caused by addition of a saturating concentration of dolichyl phosphate was greater in brain of older (3-4 weeks of age and subsequently) animals (25-fold) than in brain of younger (less than two weeks of age) animals (10-fold). This difference suggests that dolichyl phosphate may be limiting for GlcNAc-1-P transferase activity in endoplasmic reticulum of the older animals. Second, a marked (3.5-fold) increase in activity occurred over a discrete time period (3-4 weeks of postnatal life) during brain development. That this increase reflected an increase in enzyme amount rather than in catalytic efficiency was suggested by kinetic studies. Coupled with our previous demonstrations of increases in brain dolichol, dolichol kinase activity, and dolichyl phosphate levels during approximately the same developmental period (Sakakihara, Y. and Volpe, J.J., Dev. Brain Res., 14 (1984) 225-262; Volpe, J.J. et al., Dev. Brain Res., in press), the data suggest a temporally discrete period of activation of the dolichol-linked pathway to glycoproteins. Whether the pathway is regulated coordinately or sequentially is a fertile topic for future study.