We assessed the growth performance, physiological traits, and gene expressions in steers fed with dietary rumen-protected L-tryptophan (RPT) under cold environment. Eight Korean native steers were assigned to two dietary groups, no RPT (Control) and RPT (0.1% RPT supplementation on a dry matter basis), 6 wks. Maximum and minimum temperatures throughout the experiment were 6.7°C and -7.0°C, respectively. Supplementation of 0.1% RPT to a total mixed ration did not increase body weight but had positive effects of elevating average daily gain (ADG) and reducing the feed conversion ratio (FCR) at day 27 and 48. Metabolic parameter showed higher glucose level (at day 27) in the 0.1% RPT group compared to the control group. Real-time PCR analysis showed no significant differences in the expression of muscle (MYF6, MyoD, and Desmin) metabolism genes between the two groups, whereas the expression of fat (PPARγ, C/EBPα, and FABP4) metabolism genes was lower in the 0.1% RPT group than in the control group. Thus, we demonstrate that long-term (6 wks) dietary supplementation of 0.1% RPT was beneficial owing to enhanced growth performance by increasing ADG and glucose level, decreasing FCR, and maintaining homeostasis in immune responses in beef steers during cold environment.