High global uncertainties remain in water‐air CO2, CH4, and N2O fluxes from estuaries due to spatial and temporal variability and the poor predictability of the gas transfer velocity (k600). This is the first study that directly compares k600 of CO2, CH4, and N2O in an estuary with the aim to evaluate the accuracy of using a uniform k600 value for estimating water‐air fluxes. We calculated 155 k600 values from CO2, CH4, and N2O fluxes over spatial (across, along) and temporal (tidal cycle) surveys in the subtropical Maroochy estuary using the floating chamber method. Combined k600 values showed a large range over the entire estuary (0.1–198.6 cm h−1) with slightly lower k600 in the lower compared to the upper estuary. Overall, temporal variability was greater than spatial variability of k600. We found the highest variability of k600 between gas species in the lower estuary, whereas the variability was less distinct in the upper estuary. In the Maroochy estuary, k600CO2 (mean 26.4 ± 37.3 cm h−1) was mostly higher than k600CH4 (mean 10.9 ± 10.6 cm h−1) and k600N2O (mean 9.9 ± 12.3 cm h−1), likely due to chemical and enzymatic enhancements and/or microbial activity in the surface microlayer. We demonstrate that empirical k600 models intended for CO2 may not accurately predict CH4 and N2O fluxes in estuaries. Our tested k600 models predicted the measured fluxes within an uncertainty range of 5%–40% (over or underestimation), but precise flux estimates should be based on in situk600 of all three gases. Plain Language Summary: Greenhouse gas emissions from estuaries are an important component of the coastal ocean carbon cycle. However, carbon dioxide, methane, and nitrous oxide fluxes from estuaries remain uncertain at the global scale, partially because of the gas transfer velocity, a relevant parameter of the flux computation. This study is the first study, to our knowledge, that directly compares gas transfer velocities of the three greenhouse gases with each other and over spatial and temporal gradients in a subtropical estuary in Australia. We find that gas transfer velocities were gas‐specific and differed in upper, mid, and lower estuary regions. The non‐uniformity of the gas transfer velocity in heterogeneous estuaries is likely due to different chemical, physical and biological processes that drive gas transfer of the three gases at the water‐air interface. This is important to consider when estimating estuary fluxes because models for one specific gas may not accurately predict the gas transfer of the other two greenhouse gases. Key Points: k600 of CO2, CH4, and N2O were non‐uniform over spatial and temporal estuary gradients and k600CO2 mostly greater than k600CH4 and k600N2OVariability of k600 between the gas species was greater at the estuary mouth compared to the mid and upper estuary sectionsEmpirical k600CO2 models may not accurately predict CH4 and N2O fluxes because they do not account for gas‐specific non‐diffusive processes [ABSTRACT FROM AUTHOR]