A better understanding of the microalgal basic biology is still required to improve the feasibility of algal bio-products. The photosynthetic capability is one of the parameters that need further progress in research. A superior PSII activity was previously described in the green alga Neochloris oleoabundans. In this study, N. oleoabundans was grown in a glucose-supplied culture medium, in order to provide new information on the organisation and interaction of thylakoid protein complexes under mixotrophy. Fluorescence measurements suggested a strong association of light harvesting complex II (LHCII) to PSII in mixotrophic samples, confirmed by the lack of LHCII phosphorylation under growth light and the presence of PSI-PSII-LHCII megacomplexes in Blue-Native gel profile. The chloroplast ultrastructure was accordingly characterised by a higher degree of thylakoid appression compared to autotrophic microalgae. This also affected the capability of mixotrophic microalgae to avoid photodamage when exposed to high-light conditions. On the whole, it emerged that the presence of glucose affected the photosynthetic performance of mixotrophic samples, apparently limiting the dynamicity of thylakoid protein complexes. As a consequence, PSII is preserved against degradation and the PSI:PSII is lowered upon mixotrophic growth. Apparent increase in PSII photochemical activity was attributed to a down-regulated chlororespiratory electron recycling. •Mixotrophy influences the photosynthetic performance of Neochloris oleoabundans.•High FV/FM values are linked to a down-regulated chlororespiration under mixotrophy.•A stronger association of thylakoid complexes is promoted in mixotrophic cells.•Lower dynamicity of complexes allows PSII preservation under growth light.