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Semen is the main vector for HIV transmission and contains amyloid fibrils that enhance viral infection. Available microbicides that target viral components have proven largely ineffective in preventing sexual virus transmission. In this study, we establish that CLR01, a 'molecular tweezer' specific for lysine and arginine residues, inhibits the formation of infectivity-enhancing seminal amyloids and remodels preformed fibrils. Moreover, CLR01 abrogates semen-mediated enhancement of viral infection by preventing the formation of virion-amyloid complexes and by directly disrupting the membrane integrity of HIV and other enveloped viruses. We establish that CLR01 acts by binding to the target lysine and arginine residues rather than by a non-specific, colloidal mechanism. CLR01 counteracts both host factors that may be important for HIV transmission and the pathogen itself. These combined anti-amyloid and antiviral activities make CLR01 a promising topical microbicide for blocking infection by HIV and other sexually transmitted viruses.

Unlabelled: Semen enhances HIV infection in vitro, but how long it retains this activity has not been carefully examined. Immediately postejaculation, semen exists as a semisolid coagulum, which then converts to a more liquid form in a process termed liquefaction. We demonstrate that early during liquefaction, semen exhibits maximal HIV-enhancing activity that gradually declines upon further incubation. The decline in HIV-enhancing activity parallels the degradation of peptide fragments derived from the semenogelins (SEMs), the major components of the coagulum that are cleaved in a site-specific and progressive manner upon initiation of liquefaction. Because amyloid fibrils generated from SEM fragments were recently demonstrated to enhance HIV infection, we set out to determine whether any of the liquefaction-generated SEM fragments associate with the presence of HIV-enhancing activity. We identify SEM1 from amino acids 86 to 107 [SEM1(86-107)] to be a short, cationic, amyloidogenic SEM peptide that is generated early in the process of liquefaction but that, conversely, is lost during prolonged liquefaction due to the activity of serine proteases. Synthetic SEM1(86-107) amyloids directly bind HIV-1 virions and are sufficient to enhance HIV infection of permissive cells. Furthermore, endogenous seminal levels of SEM1(86-107) correlate with donor-dependent variations in viral enhancement activity, and antibodies generated against SEM1(86-107) recognize endogenous amyloids in human semen. The amyloidogenic potential of SEM1(86-107) and its virus-enhancing properties are conserved among great apes, suggesting an evolutionarily conserved function. These studies identify SEM1(86-107) to be a key, HIV-enhancing amyloid species in human semen and underscore the dynamic nature of semen's HIV-enhancing activity., Importance: Semen, the most common vehicle for HIV transmission, enhances HIV infection in vitro, but how long it retains this activity has not been investigated. Semen naturally undergoes physiological changes over time, whereby it converts from a gel-like consistency to a more liquid form. This process, termed liquefaction, is characterized at the molecular level by site-specific and progressive cleavage of SEMs, the major components of the coagulum, by seminal proteases. We demonstrate that the HIV-enhancing activity of semen gradually decreases over the course of extended liquefaction and identify a naturally occurring semenogelin-derived fragment, SEM1(86-107), whose levels correlate with virus-enhancing activity over the course of liquefaction. SEM1(86-107) amyloids are naturally present in semen, and synthetic SEM1(86-107) fibrils bind virions and are sufficient to enhance HIV infection. Therefore, by characterizing dynamic changes in the HIV-enhancing activity of semen during extended liquefaction, we identified SEM1(86-107) to be a key virus-enhancing component of human semen., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

Naturally occurring fragments of the abundant semen proteins prostatic acid phosphatase (PAP) and semenogelins form amyloid fibrils in vitro. These fibrils boost HIV infection and may play a key role in the spread of the AIDS pandemic. However, the presence of amyloid fibrils in semen remained to be demonstrated. Here, we use state of the art confocal and electron microscopy techniques for direct imaging of amyloid fibrils in human ejaculates. We detect amyloid aggregates in all semen samples and find that they partially consist of PAP fragments, interact with HIV particles and increase viral infectivity. Our results establish semen as a body fluid that naturally contains amyloid fibrils that are exploited by HIV to promote its sexual transmission.

Semen is the major vector for HIV-1 transmission. We previously isolated C-proximal fragments of the prostatic acid phosphatase (PAP) from semen which formed amyloid fibrils that potently enhanced HIV infection. Here, we used the same methodology and identified another amyloidogenic peptide. Surprisingly, this peptide is derived from an N-proximal fragment of PAP (PAP85-120) and forms, similar to the C-proximal fragments, positively charged fibrillar structures that increase virion attachment to cells. Our results provide a first example for amyloid formation by fragments of distinct regions of the same precursor and further emphasize the possible importance of amyloidogenic peptides in HIV transmission.

Many globular and natively disordered proteins can convert into amyloid fibrils. These fibrils are associated with numerous pathologies as well as with normal cellular functions, and frequently form during protein denaturation. Inhibitors of pathological amyloid fibril formation could be useful in the development of therapeutics, provided that the inhibitors were specific enough to avoid interfering with normal processes. Here we show that computer-aided, structure-based design can yield highly specific peptide inhibitors of amyloid formation. Using known atomic structures of segments of amyloid fibrils as templates, we have designed and characterized an all-D-amino-acid inhibitor of the fibril formation of the tau protein associated with Alzheimer's disease, and a non-natural L-amino-acid inhibitor of an amyloid fibril that enhances sexual transmission of human immunodeficiency virus. Our results indicate that peptides from structure-based designs can disrupt the fibril formation of full-length proteins, including those, such as tau protein, that lack fully ordered native structures. Because the inhibiting peptides have been designed on structures of dual-β-sheet 'steric zippers', the successful inhibition of amyloid fibril formation strengthens the hypothesis that amyloid spines contain steric zippers., (©2011 Macmillan Publishers Limited. All rights reserved)

Background: HIV-1 is usually transmitted in the presence of semen. We have shown that semen boosts HIV-1 infection and contains fragments of prostatic acid phosphatase (PAP) forming amyloid aggregates termed SEVI (semen-derived enhancer of viral infection) that promote virion attachment to target cells. Despite its importance for the global spread of HIV-1, however, the effect of semen on virus infection is controversial., Results: Here, we established methods allowing the meaningful analysis of semen by minimizing its cytotoxic effects and partly recapitulating the conditions encountered during sexual HIV-1 transmission. We show that semen rapidly and effectively enhances the infectivity of HIV-1, HIV-2, and SIV. This enhancement occurs independently of the viral genotype and coreceptor tropism as well as the virus producer and target cell type. Semen-mediated enhancement of HIV-1 infection was also observed under acidic pH conditions and in the presence of vaginal fluid. We further show that the potency of semen in boosting HIV-1 infection is donor dependent and correlates with the levels of SEVI., Conclusions: Our results show that semen strongly enhances the infectivity of HIV-1 and other primate lentiviruses and that SEVI contributes to this effect. Thus, SEVI may play an important role in the sexual transmission of HIV-1 and addition of SEVI inhibitors to microbicides may improve their efficacy.