The Dengue virus (DEN) belongs to the Flavivirus genus, Flaviridae Family. Nowadays, Dengue is the most important human disease caused by arbovirus around the world. There are four distinct serotypes, called DEN-1, DEN-2, DEN-3 and DEN-4. It is an enveloped virus containing a nucleocapsid that is surrounded by a membrane. The virus surface is composed of two glycoproteins: E and M. The E protein is 57-59 kDa and contains glycosilated sites. Its function is related to receptor binding and membrane fusion, during virus entry into the host cells. It is called "class II" viral fusion protein. The M protein is 8 kDa and consists of 75 amino acids. The M protein results from the cleavage of the pre-M glycoprotein during the late phase of viral maturation. Its organization consists of a short ectodomain (1-40 residues) followed by the transmembrane anchoring region (40-75 residues). Recently, M protein was correlated with apoptosis and chaperone activity on the folding of E protein. However, the exact role of M protein on virus surface is not clear. The main goal of this work is cloning, expression and re-folding of M protein, which can be useful to allow the structural studies about virus protein surface. First, the M protein ORF was amplified by PCR, and cloned into the pGEM-T easy vector followed by the cloning into the pQE30Xa and pET3-a. The clone selection was performed by the presence of blue and white colonies (pGEM-T easy vector) or by the presence of nucleotide sequence after restriction site cleavage. The data show a successful cloning and expression of M protein. In addition, the protein was purified in an unfolded conformation and refolding in the presence of some detergent micelles showing a good structural gain in relation to the protein in the absence of micelles. The structural behavior of M protein was evaluated by circular dichroism, intrinsic and extrinsic fluorescence and H¹N¹⁵ NMR. This is the first time, in which the M protein is expressed and studied in great detail. Our results on Dengue virus surface protein may provide valuable information for the design of more efficient drugs that block virus infection.