The DNMT3B de novo DNA methyltransferase (DNMT) plays a major role in 
establishing DNA methylation patterns in early mammalian development, but its 
catalytic mechanism remains poorly characterized. Here, we provide a 
comprehensive biochemical analysis of human DNMT3B function through the 
characterization of a series of site-directed DNMT3B variants associated with 
immunodeficiency, centromere instability, and facial anomalies (ICF) syndrome. 
Our data reveal several novel and important aspects of DNMT3B function. First, 
DNMT3B, unlike DNMT3A, requires a DNA cofactor in order to stably bind to 
S-adenosyl-l-methionine (SAM), suggesting that it proceeds according to an 
ordered catalytic scheme. Second, ICF mutations cause a broad spectrum of 
biochemical defects in DNMT3B function, including defects in 
homo-oligomerization, SAM binding, SAM utilization, and DNA binding. Third, all 
tested ICF mutations, including the A766P and R840Q variants, result in altered 
catalytic properties without interfering with DNMT3L-mediated stimulation; this 
indicates that DNMT3L is not involved in the pathogenesis of ICF syndrome. 
Finally, our study reveals a novel level of coupling between substrate binding, 
oligomerization, and catalysis that is likely conserved within the DNMT3 family 
of enzymes.