Vancomycin is one of the most important clinical antibiotics in the fight against infectious disease. Its biological activity relies on three aromatic cross-links, which create a cup-shaped topology and allow tight binding to nascent peptidoglycan chains. The cytochrome P450 enzymes OxyB, OxyA, and OxyC have been shown to introduce these synthetically challenging aromatic linkages. The ability to utilize the P450 enzymes in a chemo-enzymatic scheme to generate vancomycin derivatives is appealing but requires a thorough understanding of their reactivities and mechanisms. Herein, we systematically explore the scope of OxyB biocatalysis and report installation of diverse diaryl ether and biaryl cross-links with varying macrocycle sizes and compositions, when the enzyme is presented with modified vancomycin precursor peptides. The structures of the resulting products were determined using one-dimensional/two-dimensional nuclear magnetic resonance spectroscopy, high-resolution mass spectrometry (HR-MS), tandem HR-MS, and isotopic labeling, as well as ultraviolet-visible light absorption and fluorescence emission spectroscopies. An exploration of the biological activities of these alternative OxyB products surprisingly revealed antifungal properties. Taking advantage of the promiscuity of OxyB, we chemo-enzymatically generated a vancomycin aglycone variant containing an expanded macrocycle. Mechanistic implications for OxyB and future directions for creating vancomycin analogue libraries are discussed.