In the present study forty heavy metal resistant yeasts were isolated from highly metal polluted industrial wastewater of Shiekhupura and five yeast isolates showing highest metal resistance ability were identified by 18S ribotyping as Candida tropicalis, Pichia kudriawzevii, Candida sp., Rhodotorula mucilaginosa and Trichosporon asahii. The yeast isolates P. kudriavzeii, C. tropicalis, R. mucilaginosa and Candida sp. grew well and multiplied at faster rate at 30ºC with the exception of T. asahii that multiplied and divided faster at 37oC. C. tropicalis, P. kudriavzeii and R. mucilaginosa grew best at pH 7 while T. asahii and Candida sp. exhibited maximum growth at pH 6. Overall MIC of the most resistant yeast isolates was ranged from 5-30 mM and varied among yeast species. Among the heavy metals ions, Pb2+ and Cu2+ were the most tolerated heavy metals while Cr6+, Cd2+ and As3+ were the most toxic heavy metals. C. tropicalis showed a remarkable 2 fold increase in GSH content in the presence of arsenic and cadmium, one and a half (1.5) fold increase in chromium and copper stress and one half (0.5) fold under lead ions. In P. kudriavzeii a significant 2.5 fold increase in GSH level was determined by lead and copper treated cells, 2 fold increase in arsenic and cadmium while one half (0.5) fold increase was revealed in chromium. Likewise, 2.5 fold increase in GSH level was detected in R. mucilaginosa in cadmium treated cells which was almost doubled under arsenic, copper, lead and chromium treatment. Candida sp. showed enhanced GSH level in arsenic cadmium, copper and chromium treated cells while Pb2+ significantly increased GSH level by 2 fold. T. asahii cells showed remarkably elevated GSH level in cadmium and arsenic treated cells. An increase in GSH level was also determined in chromium, lead and copper treatment. Heavy metals showed remarkable increase in cysteine and non-protein thiol levels. It was determined that stressed cells had much more oxidized GSH than unstressed cells indicating the role of GSSG on the expense of GSH. Metal ions affect the activity levels of superoxide dismutase and catalase by providing defense against heavy metal ions. In R. mucilaginosa, SOD activity patterns were significantly elevated in cadmium (132%) and arsenic (119%) whereas T. asahii cells showed up-regulation 37.5% and 22.5% under cadmium and arsenic, respectively. Remarkable stimulatory effect in catalase level in red yeast, R. mucilaginosa, exposed to cadmium (237%) and arsenic (145%) was detected. Catalase activity in T. asahii cells exposed to arsenic was up-regulated (249%) and down-regulated (90%) in cadmium. Glutathione-related antioxidant activities depicted down-regulation in GST activity to 21% and 40% in arsenic and cadmium exposed yeast cells. Enhanced GR activity was detected in T. asahii exposed to cadmium (277%) and arsenic (80%) that was increased to 293% and 128% in R. mucilaginosa implying more GSSG was produced under oxidative stress. Antioxidant defenses have confirmed to play an important protective role by reducing toxicity and oxidative stress induced due to heavy metal ions. The maximum cadmium uptake potential in R. mucilaginosa determined was 30, 36, 44, 53, 60 and 65 mg/g and the cells adsorbed 3, 6, 10, 14, 19 and 22 mg/g after 2, 4, 6, 8, 10 and 12 days of growth. T. asahii cells adsorbed 3, 5, 9, 13, 17 and 20 mg/g and accumulated 26, 34, 42, 47, 52 and 61 mg/g after 2, 4, 6, 8, 10 and 12 days, respectively. Arsenic uptake by R. mucilaginosa was 27, 32, 40, 46, 53, 64 mg/g and the adsorption potential was 3, 9, 11, 16, 21 and 26 mg/g after 2, 4, 6, 8, 10 and 12 days. Arsenic uptake capacity of T. asahii was 20, 26, 31, 38, 51, 60 mg/g and the cells adsorbed 5, 8, 12, 17, 20 and 23 mg/g after 2, 4, 6, 8, 10 and 12 days from the culture medium. Highest removal potential determined in the presence of cadmium was 87% and 81% in R. mucilaginosa and T. asahii yeast cells whereas maximum arsenic removal was 90% and 83% in R. mucilaginosa and T. asahii. This metal uptake capacity of yeast isolates can be exploited in wastewater treatment strategies. Marked differences in protein banding pattern were observed in heavy metal exposed and unexposed yeast cells. All heavy metal treated C. tropicalis cultures showed a prominent 20 kDa protein band which was much reduced in untreated culture (Control). In P. kudriavzeii low molecular weight proteins were abundant in all metal-treated samples as compared to control. In cadmium, arsenic and chromium treated R. mucilaginosa cells, protein band intensity was much stronger as compared to control suggesting expression was higher under these metals. In T. asahii both cadmium and copper treatment showed low intensity 80 and 20 kDa protein bands as compared to control. A 20 kDa protein band was also appeared with increased intensity in cadmium, arsenic and chromium treated cultures of Candida sp. Two dimensional gel electrophoresis (2-DE) analyses showed the abundance of proteins on heavy metal treatment. Most protein spots identified were belong to heat shock proteins, aconitase and phosphoglycerate kinase and found to be over-expressed. Proteins belong to amino acid metabolism, protein folding and degradation pathways are principally affected under heavy metal stress conditions and may be involved in cell resistance mechanisms. These multiple-metal resistant yeast strains can be successfully used for the treatment of wastewater polluted with toxic metal ions.