Injectable and enzyme-mediated cross-linked hydrogels are promising biomedical materials. However, although poly(ethylene glycol) (PEG) is a popular basic component of synthetic hydrogels, only a few PEG-based enzymatically cross-linked hydrogels have been developed based on branched PEG. Compared with branched PEG, linear PEGs with different molecular weights are readily available and low-cost, while the poor capacity for post-polymerization modifications of linear PEG limited its application on a greater scale. Herein, a linear PEG-based analogue functionalized with multiple phenolic hydroxyl moieties, PEGDA-DTT-HPA, was designed and synthesized via Michael-type polyaddition combined with Steglich esterification. Environmentally friendly hydrogels composed of PEGDA-DTT-HPA were facilely formed under the catalysis of horseradish peroxidase (HRP) in the presence of hydrogen peroxide (H₂O₂). The gelation time and mechanical strengths of hydrogels were found to be adjusted independently by altering the concentrations of HRP and H₂O₂, respectively. The hydrogels were further demonstrated as protein drug and cell carriers using bovine serum albumin (BSA) and lentivirus-mediated LifeAct-EGFP overexpressed human mesenchymal stem cells (hMSCs-LifeAct-EGFP), respectively. The BSA-loaded hydrogel systems exhibited a sustained drug release over 3 weeks; the encapsulated hMSCs showed good viability over all time points assessed. Consequently, the current study opens new avenues for the design of PEG-based injectable hydrogels and the PEGDA-DTT-HPA hydrogel has great potential for applications in drug delivery, 3D cell culture and tissue regeneration.