Diabetes, one of the most serious and common chronic metabolic diseases affecting people worldwide in the 21st century, has become a major problem that needs to be addressed urgently. This study was designed to elucidate the anti-diabetic effect of yak yogurt-derived () SHY130 on C57BL/6J mice fed high-fat diet and streptozotocin (HFD/STZ), and the potential regulatory mechanisms involved. Mice were divided into 3 groups: normal control, diabetes, and diabetes treated with SHY130 (SHY130). Treatment with SHY130 had a regulatory effect on blood glucose and clearly ameliorated insulin resistance in T2DM mice. SHY130 inhibited the reduction in β-cell mass and α-cell proliferation in the pancreas and increased the expression of the short-chain fatty acid (SCFA) receptors GPR43 and GPR41 in the colon of T2DM mice. Furthermore, SHY130 treatment readjusted intestinal flora structure, enhanced the abundance of SCFA-producing bacteria, such as , , , and increased the levels of SCFAs in diabetic mice. In summary, SHY130 ameliorated hyperglycemia in HFD/STZ-induced diabetic mice by regulating the enteroinsular axis.