The spectral and charged characteristics of dissolved organic matter (DOM) in municipal sewage secondary effluent influenced by heavy metal ions were studied by three-dimensional fluorescence spectroscopy, ultraviolet spectrometry, etc. As the Cu(II) concentration increased, the fluorescence intensity of the DOM's fluorescence peak reduced. When the Cu(II) concentration was 100 μmol/L, the fluorescence intensity of the humic-like acid in the DOM reduced by 19.5%–27.6%. However, there was no obvious change in the fluorescence intensity when Cd(II) was added. According to the modified Stern–Volmer equation, the conditional stability constant lgK and the proportion of the fluorescent functional groups of the DOM binding with Cu(II) were 2.19–2.69 and 16%–48%, respectively. The UV253/UV203 value changed little with the additional dosage of metal ions below 10 μmol/L, which increased as the concentration of Cu(II) or Pb(II) increased from 10–100 μmol/L. As the metal concentrations increase, the absolute values of the Zeta potential and the particle sizes of the DOM basically increased. The results showed that Cu(II) had the strongest binding ability to the groups that emitted fluorescence in the secondary effluent DOM compared to Pb(II) and Cd(II).