Biology subjects

London, J.

Publications and source records attributed to London, J..

3 recordsLinked to original sources

bioRxiv2021

DNA Breaks and Gaps Target Retroviral Integration

Integration into a host genome is essential for retrovirus infection and is catalyzed by a nucleoprotein complex (Intasome) containing the viral integrase (IN) and reverse transcribed (RT) copy DNA (cDNA). Previous studies demonstrated DNA site recognition limited intasome integration. Using single molecule Forster resonance energy transfer (smFRET), we show Prototype Foamy Virus (PFV) intasomes pause at DNA strand breaks and gaps. The break/gap discontinuities are similar to base excision repair (BER) lesion-processing intermediates, which affect retrovirus integration in vivo. Pausing targeted site-directed integration at the break/gap without inducing intasome conformational alterations. An 8-oxo-guanine lesion normally processes by BER and a G/T mismatch or a +T nucleotide insertion that induce flexibility or a bend in the DNA backbone did not promote intasome pausing or targeted integration. These results suggest that repair intermediates can modulate dynamic intasome-DNA interactions which target retroviral integration.

biochemistry↗

bioRxiv2021

MutS and DNA Function as a Clamp Loader for the MutL Sliding Clamp During Mismatch Repair

DNA mismatch repair (MMR) is accomplished by highly conserved MutS and MutL homologs. MutS proteins recognize mismatch nucleotides and in the presence of ATP form a stable sliding clamp on the DNA. The MutS sliding clamp then promotes the cascade assembly of a MutL sliding clamp, which ultimately coordinates downstream mismatch excision. The MutS clamp-loader mechanics are unknown. Here we have examined a conserved positively charged cleft (PCC) located on the MutL N-terminal domain (NTD) proposed to mediate stable DNA binding events in several MMR models. We show that MutL does not bind DNA in physiological ionic conditions. Instead, the MutS sliding clamps and DNA together exploit the PCC to position the MutL NTD for clamp loading. Once in a sliding clamp form, the MutL PCC aids in UvrD helicase capture but not interactions with MutH during mismatch excision. The MutS-DNA clamp-loader progressions are significantly different from the replication clamp-loaders that attach polymerase processivity factors such as {beta}-clamp and PCNA to the DNA. These studies underlining the breadth of mechanisms for stably linking crucial genome maintenance proteins to the DNA.

biophysics↗

bioRxiv2021

Sulfated host glycan recognition by carbohydrate sulfatases of the human gut microbiota

The vast microbial community that resides in the human colon, termed the human gut microbiota, performs important roles in maintaining host health. Sulfated host glycans comprise both a major nutrient source and important colonisation factors for this community. Carbohydrate sulfatases remove sulfate groups from glycans and are essential in many bacteria for the utilisation of sulfated host glycans. Additionally, carbohydrate sulfatases are also implicated in numerous host diseases, but remain some of the most understudied carbohydrate active enzymes to date, especially at the structural and molecular level. In this work, we analyse 7 carbohydrate sulfatases, spanning 4 subfamilies, from the human gut symbiont Bacteroides thetaiotaomicron, a major utiliser of sulfated host glycans, correlating structural and functional data with phylogenetic and environmental analyses. Together, these data begin to fill the knowledge gaps in how carbohydrate sulfatases orchestrate sulfated glycan metabolism within their environment.

biochemistry↗