Conventional views associate microbial biofilm with demineralization in root caries (RC) onset, while research on their collagenases role in the breakdown of collagen matrix has been sporadically developed, primarily in vitro. Recent discoveries, however, reveal proteolytic bacteria enrichment, specially Porphyromonas and other periodontitis-associated bacteria in subgingivally extended lesions, suggesting a potential role in RC by the catabolism of dentin organic matrix. Moreover, genes encoding proteases and bacterial collagenases, including the U32 family collagenases, were found to be overexpressed in both coronal and root dentinal caries. Despite these advancements, to prove microbial collagenolytic proteases' definitive role in RC remains a significant challenge. A more thorough investigation is warranted to explore the potential of anti-collagenolytic agents in modulating biofilm metabolic processes or inhibiting/reducing the size of RC lesions. Prospective treatments targeting collagenases and promoting biomodification through collagen fibril cross-linking show promise for RC prevention and management. However, these studies are currently in the in vitro phase, necessitating additional research to translate findings into clinical applications. This is a comprehensive state-of-the-art review aimed to explore contributing factors to the formation of RC lesions, particularly focusing on collagen degradation in root tissues by microbial collagenases.