Ubiquitin-like proteins constitute a diverse group of post-translational modifiers that share structural and functional similarities with Ubiquitin, but play distinct roles in cellular signaling. The ubiquitin-like protein FUBI acts as a positive regulator of apoptosis in cancers, where its down-regulation is significantly associated with poor prognosis. Despite the clinical and biological relevance of FUBI-related functions, the intricate regulatory mechanisms orchestrated by FUBI remain elusive. To address this knowledge gap, we developed a linear synthetic platform to generate FUBI-based chemical tools. This platform enables the site-specific incorporation of unnatural building blocks and the introduction of fluorophores, tags, reactive warheads through N- and C-terminal labeling strategies. Using this platform, we created activity-based probes (ABPs) for FUBI conjugation and deconjugation enzymes, validating them in cell lysate-based assays and proteomics. Additionally, we synthesized K25 triazole-linked Di-FUBI as bioisosteric uncleavable mimic of native Di-FUBI to study the proteolytic cleavage of FUBI chains. Collectively, these ABPs demonstrate the versatility and specificity of our synthetic FUBI platform, advancing the characterization of FUBI-related enzymes and decoding their roles in cellular processes at the molecular level.