Background: Viral evolution of HIV-1 is dynamic and moving towards a higher order of replicative fitness. Results: HIV-1 subtype C acquires an extra (4th) NF-B site to achieve a higher degree of transcription and in turn enhances its replicative fitness and preponderance. Conclusion: Subtype C with an extra NF-B site adopts a novel strategy of strengthening its promoter to gain fitness. Significance: Learning how the new strains could impact viral prevalence, pathogenesis, and disease management strategies is critical.

The outer domain (OD) of the HIV-1 envelope glycoprotein gp120 is an important target for vaccine design as it contains a number of conserved epitopes, including a large fraction of the CD4 binding site. Attempts to design OD-based immunogens in the past have met with little success. We report the design and characterization of an Escherichia coli-expressed OD-based immunogen (OD EC ), based on the sequence of the HxBc2 strain. The OD EC -designed immunogen lacks the variable loops V1V2 and V3 and incorporates 11 designed mutations at the interface of the inner and the outer domains of gp120. Biophysical studies showed that OD EC is folded and protease-resistant, whereas OD EC lacking the designed mutations is highly aggregation-prone. In contrast to previously characterized OD constructs, OD EC bound CD4 and the broadly neutralizing antibody b12 but not the non-neutralizing antibodies b6 and F105. Upon immunization in rabbits, OD EC was highly immunogenic, and the sera showed measurable neutralization for four subtype B and one subtype C virus including two b12-resistant viruses. In contrast, sera from rabbits immunized with gp120 did not neutralize any of the viruses. OD EC is the first example of a gp120 fragment-based immunogen that yields significant neutralizing antibodies.The envelope glycoprotein of human immunodeficiency virus 1 (HIV-1) is composed of two polypeptide chains, gp120 and gp41, that are present on the surface of the virus as a trimer of heterodimers (1). As these are the primary viral components exposed to the external environment, they present the single best target for neutralizing antibodies. gp41 mainly contains the fusion machinery and a large part of the membrane proximal region and is transiently exposed during the fusion process (2, 3). gp120 remains largely exposed on the virus (4). However, the virus has evolved various mechanisms to evade the humoral immune response toward gp120. A very high rate of mutation, large conformational flexibility (5), and the extensive glycosylation of the surface (6, 7) are some of the important ways by which the virus has evaded a neutralizing antibody response (8). As a result, when recombinant gp120 is used as an immunogen, the antibodies generated are often directed to immunodominant epitopes present in variable loops and have limited breadth of neutralization (9 -13). Core gp120 lacking the variable loops is poorly immunogenic, in part because of surface coverage by glycans (2, 14). It was recently shown that demannosylated full-length gp120 is much more immunogenic than wild-type gp120 in mouse immunization studies (15).The structure of gp120, as seen from the crystal structure of gp120 complexed to CD4 and antibody 17b (PDB ID 1G9M) (16) reveals that the molecule can be subdivided into three distinct parts: the inner domain, the outer domain, and the bridging sheet. On monomeric gp120, the inner domain is well exposed and has little coverage by glycans (2). However, this domain is likely to be involved in interactions with gp41 (17) and is pro...

Linker histone H1 plays an essential role in chromatin organization. Proper deposition of linker histone H1 as well as its removal is essential for chromatin dynamics and function. Linker histone chaperones perform this important task during chromatin assembly and other DNA-templated phenomena in the cell. Our in vitro data show that the multifunctional histone chaperone NPM1 interacts with linker histone H1 through its first acidic stretch (residues 120-132). Association of NPM1 with linker histone H1 was also observed in cells in culture. NPM1 exhibited remarkable linker histone H1 chaperone activity, as it was able to efficiently deposit histone H1 onto dinucleosomal templates. Overexpression of NPM1 reduced the histone H1 occupancy on the chromatinized template of HIV-1 LTR in TZM-bl cells, which led to enhanced Tat-mediated transactivation. These data identify NPM1 as an important member of the linker histone chaperone family in humans.

Protein post-translational modifications (PTMs) play important roles in the control of various biological processes including protein–protein interactions, epigenetics and cell cycle regulation. Mass spectrometry-based proteomics approaches enable comprehensive identification and quantitation of numerous types of PTMs. However, the analysis of PTMs is complicated by the presence of indistinguishable co-eluting isomeric peptides that result in composite spectra with overlapping features that prevent the identification of individual components. In this study, we present Iso-PeptidAce, a novel software tool that enables deconvolution of composite MS/MS spectra of isomeric peptides based on features associated with their characteristic fragment ion patterns. We benchmark Iso-PeptidAce using dilution series prepared from mixtures of known amounts of synthetic acetylated isomers. We also demonstrate its applicability to different biological problems such as the identification of site-specific acetylation patterns in histones bound to chromatin assembly factor-1 and profiling of histone acetylation in cells treated with different classes of HDAC inhibitors.