Data from: Identification of allosteric disulphides from labile bonds in X-ray structures

Protein disulfide bonds link pairs of cysteine sulfur atoms and are either structural or functional motifs. The allosteric disulfides control the function of the protein in which they reside when cleaved or formed. Here, we identify potential allosteric disulfides in all Protein Data Bank X-ray structures from bonds that are present in some molecules of a protein crystal but absent in others, or present in some structures of a protein but absent in others. We reasoned that the labile nature of these disulfides signifies a propensity for cleavage and so possible allosteric regulation of the protein in which the bond resides. A total of 511 labile disulfide bonds were identified. The labile disulfides are more stressed than the average bond, being characterized by high average torsional strain and stretching of the sulfur–sulfur bond and neighbouring bond angles. This pre-stress likely underpins their susceptibility to cleavage. The coagulation, complement and oxygen-sensing hypoxia inducible factor-1 pathways, which are known or have been suggested to be regulated by allosteric disulfides, are enriched in proteins containing labile disulfides. The identification of labile disulfide bonds will facilitate the study of this post-translational modification

Table S2

Excel spreadsheet of labile disulphide bonds present in some molecules of a protein crystal but absent in others (same PDB, sheet 1), or present in some structures of a protein but absent in others (different PDB, sheet 2)

Table S1

Excel spreadsheet of unique disulphides in a culled set of X-ray structures described by G. Wang and R. Dunbrack, Jr. (file pdbaanr)

Supplementary material from Identification of allosteric disulfides from labile bonds in X-ray structures

Table S3, Figures S1-S

Table S1 from Identification of allosteric disulfides from labile bonds in X-ray structures

Excel spreadsheet of unique disulphides in a culled set of X-ray structures described by G. Wang and R. Dunbrack, Jr. (file pdbaanr)

Table S2 from Identification of allosteric disulfides from labile bonds in X-ray structures

Excel spreadsheet of labile disulphide bonds present in some molecules of a protein crystal but absent in others (same PDB, sheet 1), or present in some structures of a protein but absent in others (different PDB, sheet 2)

Development of sensitive ddPCR assays to reliably quantify the proviral DNA reservoir in all common circulating HIV subtypes and recombinant forms

Introduction: The latent reservoir is the main barrier on the road to HIV cure, and clinical approaches towards eradication are often evaluated by their effect on proviral DNA. To ensure inclusiveness and representativeness in HIV cure studies, proviral DNA quantification assays that are able to detect all common circulating HIV clades are urgently needed. Here, three HIV DNA assays targeting three different genomic regions were evaluated for their sensitivity and subtype-tolerance using digital PCR. Methods: A subtype-B-specific assay targeting gag (GAG) and two assays targeting conserved sequences in ltr and pol (LTR and JO) were assessed for their sensitivity and subtype-tolerance in digital PCR (Bio-Rad QX200), using a panel of serially diluted subtype reference plasmids as well as a panel of clinical isolates. Both panels represent subtypes A, B, C, D, F, G and circulating recombinant forms (CRFs) AE and AG, which together are responsible for 94% of HIV infections worldwide. Results: HIV subtype was observed to greatly affect HIV DNA quantification results. Robust regression analysis of the serially diluted plasmid panel showed that the GAG assay was only able to linearly quantify subtype B, D and G isolates (4/13 reference plasmids, average R2 = 0.99), whereas LTR and JO were able to quantify all tested isolates (13/13 reference plasmids, respective average R2 = 0.99 and 0.98). In the clinical isolates panel, isolates were considered detectable if all replicates produced a positive result. The GAG assay could detect HIV DNA in four out of five subtype B and one out of two subtype D isolates, whereas the LTR and JO assays detected HIV DNA in all twenty-nine tested isolates. LTR and JO results were found to be equally precise but more precise than GAG. Conclusions: The results demonstrate the need for a careful validation of proviral reservoir quantification assays prior to investigations into non-B subtype reservoirs. The LTR and JO assays can sensitively and reliably quantify HIV DNA in a panel that represents the worldwide most prevalent subtypes and CRFs (A, B, C, D, AE, F, G and AG), justifying their application in future trials aimed at global HIV cure