This work reports a thermoresponsive multifunctional wound dressing hydrogel based on ABA triblock copolymers synthesized via reversible addition fragmentation chain transfer (RAFT) polymerization. The inner B block consists of a positively-charged hydrolysable betaine ester loaded with an antimicrobial drug as its counter ion and the B block is flanked by two outer A blocks of thermoresponsive poly (N-isopropylacrylamide) (PNIPAM). A solution containing the triblock copolymers can be applied to wound sites and immediately turns into a physical gel at the body temperature. This wound dressing can reduce the risk of wound infection by releasing small-molecular-weight antimicrobial drug and facilitate the attachment of mammalian cells during tissue regeneration through its positive surface charge. The cationic betaine ester can then hydrolyze at the wound site to its zwitterionic form, which is known to be biocompatible and nonsticky. The thermoresponsive in situ gelation feature along with controlled drug release, enhanced tissue–hydrogel interactions as well as long-term biocompatibility make this hydrogel a very promising material for antimicrobial wound dressing applications.