The businesses in the former Debenhams store will move out after Saturday.
The closure comes amid news of the demolition of the former Postings shopping center which sits behind it on Hill Street.
Both buildings are owned by Wakefield businessman Tahir Ali.
The department store runs under Hill Street and will be directly affected by major plans to level the old center and clear land for the proposed housing development.
Staff were briefed on the plan a few months ago.
Pupils at Fife School have moved out as major works begin to create more space
Mr Ali set up Fife Department Store in November 2021.
It came after the 15-17 former leaseholder collapsed in chaos with many concessions claiming they had been left out.
15-17 was mired in claims from shopkeepers and contractors at its other UK stores who weren’t getting paid – reports say the Kirkcaldy branch had a similar situation.
The move proved popular with buyers and allowed businesses to continue to have a presence in the High Street.
The boutique will close to allow the adjacent New Look to continue its activities.
The two units are linked – they once housed the old Littlewoods store – and both have storage facilities and staff rooms that run under Hill Street below where Tesco used to sit.
The complex nature of the proposed demolition also led Mr Ali to buy a number of High Street buildings, including Bank of Scotland, Savers and Bob & Berts.
Not all businesses are affected by this week’s announcements.