These days many people around the world suffer from obesity, insulin resistance, and diabetes type II. Sugar, the most favorite sweetener, is a cause of these diseases due to high calorie level. Therefore, finding healthy and financially reasonable replacements for sugar and artificial sweeteners has increasingly become important and the demand for non-calorigenic protein-based sweeteners with favorable taste properties is high. So far, eight sweet proteins including Brazzein have been found in the African plant of Pentadiplandra Brazeanna. Brazzein has become an important sweetener protein due to sweetness, heat stability, minimum molecular weight, and having the characteristics of carbon hydrates. In the present work we aim to produce Brazzein in E.coli as fusion protein with His.Tag. Threfore the sequence of the gene encoding Brazzein was coloned into pET28a. The resulted constructs pET28a- braz were transformed to E.coli , strain Rosseta (DE3). Following the induction with isopropyl-?-D-1-thiogalactopyranoside, the protein His-Breaz was expressed in E. coli . However a large percent of protein was found in insoluble fraction. To solve the solubility of protein we decided to express the protein as carboxyl-terminal extensions of glutathione-S-transferase (GST-tag). The analysis of total protein as well as soluble and insoluble fractions of E. coli revealed that the presence of GST-tag resulted in 40 % increase in solubility. Both of His-Braz and GST-Braz were purified using affinity chromatography. We found that the concentration (based on molarity) and the final yield of GST.Braz (the produced pure protein from 1 L culture) are 512 and 3200 times more than His.Braz. Because of the limited effect of N-terminal tags on the sweetness of Brazzein, we tried to remove the GST-tag using enterokinase enzyme. The analysis of digested product on Tricine SDS-PAGE revealed that the enzymatic digestion was effective. However the challenge of enzyme cost in production of large scale Brazzein is a serious problem in using GST. braz system Therefore the production of Brazzein in eukaryote system like Yeast Saccharomyces cerevisiae was another step in this project. To this end the gene encoding Brazzein was cloned in a vector with GPD promoter. The construct GPD-Braz was transformed into 5D strain of the yeast. The expression of Brazzein was confirmed in Saccharomyces cerevisiae after analysis of total protein on Tricine SDS-PAGE. However the solubility of protein in this system and the sweetness of protein is still required to be checked in the future. Keywords: Sweet protein, Brazzein, fusion Tag GST, Enterokinase enzyme Tricine-SDS-PAGE