Four batch experiments of hydrolysis and acidification were carried out to investigate the distributions of proteins (PN) and polysaccharides (PS) in the sludge, the PN/PS ratio, the particle sizes, and their relationship with sludge dewaterability (as determined by capillary suction time, CST). The sludge flocs were stratified through centrifugation- and ultrasound-based method into four fractions: (1) slime, (2) loosely bound extracellular polymeric substances (LB-EPS), (3) tightly bound EPS (TB-EPS), and (4) pellet. The results showed that PN was mainly partitioned in the pellet (80.7%) and TB-EPS (9.6%) fractions, while PS distributed evenly in the four fractions. During hydrolysis and acidification, PN was transferred from the pellet and TB-EPS fractions to the slime fraction, but PS had no significant transfer trends. The mean particle sizes of the sludge flocs decreased with hydrolysis and acidification. The pH had a more significant influence on the dewaterability of sludge flocs than temperature. Sludge dewaterability during hydrolysis and acidification processes greatly deteriorated from 9.7 s at raw sludge to 340-450 s under alkaline conditions. However, it was just slightly increased under acidic conditions. Further investigation suggested that CST was affected by soluble PN, soluble PN/PS, and particle sizes of sludge flocs, but was affected slightly by total PIN, PS, or PN/PS in the whole sludge flocs and other fractions (except slime).