A clamp-like biohybrid catalyst for DNA oxidation

Abstract

In processive catalysis, a catalyst binds to a substrate and remains bound as it performs several consecutive reactions, as exemplified by DNA polymerases. Processivity is essential in nature and is often mediated by a clamp-like structure that physically tethers the catalyst to its (polymeric) template. In the case of the bacteriophage T4 replisome, a dedicated clamp protein acts as a processivity mediator by encircling DNA and subsequently recruiting its polymerase. Here we use this DNA-binding protein to construct a biohybrid catalyst. Conjugation of the clamp protein to a chemical catalyst with sequence-specific oxidation behaviour formed a catalytic clamp that can be loaded onto a DNA plasmid. The catalytic activity of the biohybrid catalyst was visualized using a procedure based on an atomic force microscopy method that detects and spatially locates oxidized sites in DNA. Varying the experimental conditions enabled switching between processive and distributive catalysis and influencing the sliding direction of this rotaxane-like catalyst.

Light-powered autonomous and directional molecular motion of a dissipative self-assembling system

“…Although the system as such cannot be used to perform work in a bulk solution 37 , it provides a viable route to the construction of rotary motors based on catenanes 39 or linear motors 40 based on polymeric motifs 9,11 , which are capable of moving-and possibly transporting cargo-over long distances. It can also be regarded as a precursor of artificial molecular pumps 38 capable of generating concentration gradients across membranes [41][42][43][44] .…”

“…4a) confirms the validity of our mechanistic hypotheses. A few interesting facts can be extracted from the simulation to assess the performance of the cycle (Supplementary equations (11) to (13)). The cycling rate is 1.7 × 10 −10 M s −1 and the efficiency is 2.3 × 10 −3 cycles per photon; that is, about 430 photons need to be absorbed on average to complete a cycle.…”

Processive Catalysis

“…applied this strategy to an artificial DNA‐nicking catalyst (a manganese tripyridyl porphyrin) by conjugating it to the T4 clamp protein discussed earlier (Figure 8). 29 Apart from its new catalytic ability, the resulting hybrid catalyst behaved just like the natural clamp: it could interact with clamp loader proteins which actively load the clamp on specific loader sites in DNA, and it could then slide over the DNA. The catalytic ability of the conjugated porphyrin, which intercalates and then nicks DNA at AAA sequences, combined with the clamp’s sliding behavior to form a processive catalyst.…”

Wholly Synthetic Molecular Machines

“…[138] Nolte [139][140][141][142] has designed ar otaxanet hat mimics the action of these processive enzymes to catalyze many reactions, one after another in ar ow (Figure 13). The synthesis of double-helical DNA, as part of the process of replication, requires that the enzyme DNA polymerase attaches itself to as ingle DNA polymer chain prior to identifying the complimentary nucleotide andc atalyzing the formation of ap hosphate ester bond before movingt o the next nucleotide.…”

“…Many processive enzymes of this kind have at oroidal shape which allows them to encircle the biopolymer while moving along its length. [138] Nolte [139][140][141][142] has designed ar otaxanet hat mimics the action of these processive enzymes to catalyze many reactions, one after another in ar ow (Figure 13). His catalyst consists of as ubstrate-binding cavity within at riangulars tructure incorporating am anganese(III) porphyrin complexc apable of oxidizing olefins to expoxides inside its toroidal cavity.W hen the catalyst threads onto ap olybutadiene strand,i tc an epoxidize the double bonds along the polymer chain in ap rocessive manner.…”