As part of the deal the 28-year-old will not be able to play for the U's when they visit Leeds on Saturday.
U's manager Aidy Boothroyd told the club's website: "I'm pleased to be bringing in an experienced player, who's played at a much higher level."
Prutton began his career at Nottingham Forest, before earning a move to the Saints in 2003.
He becomes Colchester's sixth signing of the January transfer window, following in the footsteps of Kayode Odejayi, John Joe O'Toole, Ian Henderson, Morten Knudsen and Christian Ribeiro.
Source: BBC Sport