Soccer is the most popular sport worldwide. Despite its global acclaim, scientific studies of soccer have tended to focus on tactics and techniques, thereby neglecting the physical and physiological profile of the players. Therefore, the purpose of this study was to examine physical and anthropometric characteristics of male South African university soccer players. Twenty-seven male soccer players aged 19 to 24 (mean age: 22.1 years; s = 1.5 years) volunteered to participate in the study. The results showed that goalkeepers (77.5 ± 9.7 kg) and defenders (68.2 ± 6.5 kg) were the heaviest compared to players in other playing positions. The goalkeepers also had the highest percentage of body fat (11.3 ± 2.3%), in contrast to midfielders who had the lowest body fat content (9.1 ± 0.9%). With regard to flexibility, defenders (45.1 ± 4.9 cm) and midfielders (45.9 ± 5.4 cm) performed better than goalkeepers (37.1 ± 4.3 cm) and strikers (40.1 ± 3.4 cm). Midfielders (57.2 ± 3.1 ml1·kg−1·min1) and defenders (56.1 ± 5.1 ml1·kg−1·min1) had significantly higher values of maximal oxygen uptake (VO2max) than goalkeepers (47.9 ± 0.2 ml−1·kg−1·min−1) and strikers (49.8 ± 6.2 ml−1·kg−1·min−1). No significant (p > 0.05) differences were observed for all other variables, with the exception of body height, body mass, and VO2max. It was therefore concluded that sports scientists and coaches should tailor conditioning programmes in soccer according to players’ positions in view of the implications for successful performance.