Biology subjects

Andree, G. A.

Publications and source records attributed to Andree, G. A..

2 recordsLinked to original sources

bioRxiv2025

Cysteine availability tunes ubiquitin signaling via inverse stability of LRRC58 E3 ligase and its substrate CDO1

Cellular responses to amino acid fluctuations often hinge on ubiquitin-mediated control of metabolic enzymes, yet the underlying E3 ligase pathways remain incompletely defined. Using quantitative proteomics and active cullin-RING ligase (CRL) profiling, we identify LRRC58 as a cysteine-responsive substrate receptor whose stability increases sharply under cysteine starvation. Proteomics reveals an inverse relationship between LRRC58 and the metabolic enzyme cysteine dioxygenase 1 (CDO1), suggesting a cysteine-linked regulatory axis. Biochemical reconstitution and cryo-EM structures show that LRRC58 forms an active CUL2-or CUL5-based CRL that selectively positions CDO1 for ubiquitylation at Lys8. By contrast, targeted protein degradation via the VHL-based CRL is achieved by recruitment of a distinct surface of CDO1, and more promiscuous ubiquitylation independent of Lys8. Together, our proteomics-guided discovery pipeline, cellular stability studies, and structural analyses uncover a metabolically-tuned LRRC58-CDO1 pathway that links cysteine availability to selective proteasomal turnover, reveals principles of metabolite-regulated CRL activity, and showcases mechanisms distinguishing endogenous and targeted protein degradation.

biochemistry↗

bioRxiv2021

How ATP and dATP act as molecular switches to regulate enzymatic activity in the prototypic bacterial class Ia ribonucleotide reductase

Class Ia ribonucleotide reductases (RNRs) are subject to allosteric regulation to maintain the appropriate deoxyribonucleotide levels for accurate DNA biosynthesis and repair. RNR activity requires a precise alignment of its 2 and {beta}2 subunits such that a catalytically-essential radical species is transferred from {beta}2 to 2. In E. coli, when too many deoxyribonucleotides are produced, dATP binding to RNR generates an inactive 4{beta}4 state in which {beta}2 and 2 are separated, preventing radical transfer. ATP binding breaks the -{beta} interface, freeing {beta}2 and restoring activity. Here we investigate the molecular basis for allosteric activity regulation in the prototypic E. coli class Ia RNR. Through the determination of six crystal structures we are able to establish how dATP binding creates a binding pocket for {beta} on that traps {beta}2 in the inactive 4{beta}4 state. These structural snapshots also reveal the numerous ATP-induced conformational rearrangements that are responsible for freeing {beta}2. We further discover, and validate through binding and mutagenesis studies, a previously unknown nucleotide binding site on the subunit that is crucial for the ability of ATP to dismantle the inactive 4{beta}4 state. These findings have implications for the design of allosteric inhibitors for bacterial RNRs.

biochemistry↗