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bioRxiv · 10.1101/2022.09.24.509349

Regulation of the Phage Lambda Viral Packaging Motor's Grip and DNA End-clamp Mechanism

Rawson, B.·Yang, Q.·Catalano, C. E.·Smith, D. E.
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Abstract

Many dsDNA viruses utilize ATP-powered "terminase" motors to package their genomes into procapsid shells. Here we use a single-molecule DNA grip/slip assay with rapid solution exchange to probe effects of nucleotide binding/dissociation in phage lambda motors containing both the large (TerL) and small (TerS) terminase subunits. Both subunits are required for packaging in vivo, but for some viruses (e.g., phages T4, HK97) packaging can be measured in vitro with only the catalytic TerL subunit. TerS facilitates initiation of packaging in vivo, but it has remained unclear if it plays any role during translocation. Surprisingly we measure frequent DNA gripping and high motor-DNA friction even in the absence of nucleotide. Such behavior was not observed in phage T4 motors containing only TerL, for which motor-DNA interactions were measured to be much weaker and significant gripping and friction was only observed with nucleotide present. For the lambda TerL/TerS holoenzyme, binding of nucleotide (ATP analogs or ADP) further increases gripping and friction, indicating there are both nucleotide independent and dependent interactions. Our findings suggest that TerS plays an important role in motor processivity, and that ATP-independent DNA gripping explains pausing observed during lambda packaging. We propose TerS acts as a "sliding clamp" to limit back slipping when TerL loses grip. Additionally, we show that the lambda packaging complex has a "DNA end clamp" mechanism that prevents the viral genome from completely exiting the capsid once packaging has initiated.

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Rawson, B., Yang, Q., Catalano, C. E., Smith, D. E.. 2022-09-25. Regulation of the Phage Lambda Viral Packaging Motor's Grip and DNA End-clamp Mechanism. https://doi.org/10.1101/2022.09.24.509349

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