Richard Ismail, known as Moody, died on Sunday but had been in hospital since the incident on November 11, 2012, outside Lloyds bar in Division Street.
Police arrested a 19-year-old man and a 24-year-old man on suspicion of assault at the time of the incident last year but no-one has been charged over the attack.
Mr Ismail, from Sheffield, is thought to have been on a night out with his family at the time of the incident and sustained a head injury in the attack.
A murder investigation is now underway and South Yorkshire Police are renewing their appeal for witnesses and information.
Detective Chief Inspector Natalie Shaw, who is leading the appeal, said: “We appreciate that this incident occurred over a year ago, but we are sure that there are witnesses out there that haven’t yet come forward.
“People know who did this, and they need to have the courage to tell us what happened. Richard and his family deserve this at the very least.”
Leeds United supporters and the club’s striker Ross McCormack have been paying tribute to Mr Ismail since his death over the weekend.
Anyone with information should contact South Yorkshire Police on 101 quoting incident number 67 of 11 November 2012.