Flavivirus capsid (C) protein is a key structural component of virus particles. The non-structural role of C protein in the pathogenesis of arthropod-borne flaviviruses is not clearly deciphered. This study showed that West Nile virus (WNV) and dengue virus (DENV) utilized C protein to reduce human Sec3p (hSec3p) levels at post-transcriptional level through activation of chymotrypsin-like proteolytic function of 20S proteasome. Mutagenesis studies confirmed amino acids 14, 109–114 of WNV C protein and 13, 102–107 of DENV C protein played an important role in activating the proteolytic function of 20S proteasome. Amino acid residues at 14 (WNV) and 13 (DENV) of C protein were important for C protein-hSec3p binding and physical interaction between C protein and hSec3p was essential to execute hSec3p degradation. Degradation motif required to degrade hSec3p resided between amino acid residues 109–114 of WNV C protein and 102–107 of DENV C protein. Proteasomes, hSec3p binding motif and degradation motif on C protein must be intact for efficient flavivirus production. Clinical isolates of DENV showed more pronounced effect in manipulating the proteasomes and reducing hSec3p levels. This study portrayed the non-structural function of C protein that helped the flavivirus to nullify the antiviral activity of hSec3p by accelerating its degradation and facilitating efficient binding of elongation factor 1α with flaviviral RNA genome.