The maximum rate of carboxylation (Vcmax), a key parameter indicating photosynthetic capacity, is commonly fixed as a constant by vegetation types and/or varies according to empirical scaling functions in Earth system models (ESMs). As such, the setting of Vcmax results in uncertainties of estimated carbon assimilation. It is known that the coupling between leaf chlorophyll and Rubisco (ribulose-1,5-biphosphate carboxylase-oxygenase) contents can be applied to estimate Vcmax. However, how this coupling is affected by environmental changes and varies among plant functional types (PFTs) has not been well investigated yet. The effect of varying coupling between chlorophyll and Rubisco contents on the estimation of Vcmax is still not clear. In this study, we compiled data from 76 previous studies to investigate the coupling between Chlorophyll (Chl) and Rubisco (Rub), in different PFTs and under different environmental conditions. We also assessed the ability of a Rub-based semi-mechanistic model to estimate Vcmax normalized to 25 °C (Vcmax25) based on the Rub–Chl relationship. Our results revealed strong, linear Rub-Chl relationships for different PFTs (R 2 = 0.73, 0.67, 0.54 and 0.72 for forest, crop, grass and shrub, and C4 plants, respectively). The Rub–Chl slope of natural C3 PFTs was consistent and significantly different from those of crops and C4 plants. A meta-analysis indicated that reduced light intensity, elevated CO2, and nitrogen addition strongly altered Rub/Chl. A semi-mechanistic model based on PFT-specific Rub–Chl relationships was able to estimate Vcmax25 with high confidence. Our findings have important implications for improving global carbon cycle modeling by ESMs through the improved parameterization of Vcmax25 using remotely sensed Chl content.