Optical tweezers have become a versatile tool in the biological sciences. Combined with various types of optical microscopy, they are being successfully used to discover the fundamental mechanism of biological processes. Recently, the study of proteins acting on DNA was aggressively undertaken at the single-molecule level. Here, we review the most recent studies which have revealed the dynamic behavior of individual protein molecules at work on DNA, providing detailed mechanistic insight that could not be revealed, at least not easily, using bulk-phase or ensemble approaches.