Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco; EC 4.1.1.39) is one of the key enzymes involved in assimilation of CO2 in chloroplasts. Phylloplane microfungi and their metabolites have been reported to affect the physiology of host plants, particularly, their photosynthesis. However, information is lacking on the effect of these microflora on the physiology of chloroplasts. The current study emphasized the impact of two dominant phylloplane fungi, Aspergillus niger and Fusarium oxysporum, on activity of Rubisco in tomato chloroplasts. Ergosterol, which is a component of only fungal cell membranes and is not synthesized by plants, have been demonstrated to elicit activity of Rubisco. In the present study, it was demonstrated through in silico, in vitro, and in vivo approaches. Results demonstrated that the fungal metabolites, which contained ergosterol, could double Rubisco activity. Maximum carboxylation rate of Rubisco increased also in ergosterol-treated plants. Michaelis-Menten constant of Rubisco was also slightly affected. Ergosterol was found also to influence and enhance the binding of CO2 and ribulose-1,5-bisphosphate to Rubisco. Therefore we can postulate that the physiology of the chloroplast is probably influenced by phylloplane microfungi., J. Mitra, P. Narad, P. K. Paul., and Obsahuje bibliografii