This study aimed to assess the health literacy (HL) related knowledge, attitude, perceived barriers, and practice among primary care doctors (PCDs) in Malaysia, and to determine the factors associated with HL-related practice. A cross-sectional study was conducted using an online questionnaire. Sociodemographic and work-related details were collected. HL-related knowledge, attitude, perceived barriers, and practice were assessed. Descriptive and inferential analyses using linear regression were performed. 373 PCDs were included in the study with a mean (SD) age of 37.9 (8.1) years old. The mean (SD) HL-related knowledge, attitude, and practice scores were 6.89 (1.27), 36.33 (7.04), and 30.14 (4.7), respectively. 90.9% of the participants had good HL-related knowledge scores, and 89.5% had positive HL-related attitude. More than 80% of participants found that "time constraint to implement health literacy screening" and "lack of human resources to administer HL screening tools in their settings" were among the barriers for them to implement HL practices. PCDs of Chinese and other ethnicities had lower HL-related practice scores compared to those of Malay ethnicity (adjusted b = - 1.74; 95% CI - 2.93, - 0.54, and - 2.94; 95% CI - 5.27, - 0.60, respectively). PCDs who had heard of the term "health literacy" were associated with higher HL-related practice scores (adjusted b = 2.32; 95% CI 1.17, 3.47). Age (adjusted b = 0.10; 95% CI 0.04, 0.16) had significant linear positive relationship with HL-related practice. In conclusion, the HL-related knowledge, attitude, and practice among PCDs in Malaysia were at an acceptable level. Along with educating PCDs on HL, the perceived barriers identified need to be addressed to improve the HL-related practice and ultimately patient care.