The development of multifunctional semi-conductor nano-enzymes has attained significant interest recently, due to their ease in fabrication, cost-effectiveness, and wide range of applications. In this work, we report the synthesis of ammine-functionalized Mn3O4 quantum dots (Mn3O4 QDs) with excellent luminescence properties. These QDs have been developed via the reduction of permanganate using L-cysteine through a simple, facile one-step bottom-up approach. The steady-state enzyme kinetic studies revealed the superior Catechol-Oxidase mimic activity of the Mn3O4 QDs with a Michaelis-Menten constant (Km) of 133.19 µM, and maximum reaction velocity (Vmax) of 69.40 µMs−1. Due to this superior enzymatic activity, dopamine (DA) gets easily oxidized into DA-quinone and eventually polymerized to form polydopamine. Schiff base/Michael addition reaction between nucleophilic ammine on the surface of QDs and the aromatic chain makes the probe and analyte come closer and leads to the formation of a charge transfer complex, resulting in fluorescence quenching of QDs. This fluorescence quenching has been quantified to develop a turn-off sensor for DA. The sensing characteristics are highly selective and sensitive towards DA with a detection limit of 18 nM. The synthesised Mn3O4 QDs with superior enzyme-mimicking properties have tremendous potential as fluorescent probes in imaging and health care applications.