The extracellular processing of HIV‐1 envelope glycoprotein gp160 by human plasmin

Okumura, Yuushi; Yano, Mihiro; Murakami, Meiko; Mori, Sachie; Towatari, Takae; Kido, Hiroshi

doi:10.1016/s0014-5793(98)01612-3

Cited by 17 publications

(11 citation statements)

References 24 publications

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“…1A). This is consistent with the previous report on gp160 cleavage by plasmin (61). However, when a Western blot of the 16-h plasmin digest was probed with the gp120-specific MAb, B12, it was clear that plasmin also digests gp120 into fragments, one of which is about 70 kDa (Fig.…”

Section: Resultssupporting

confidence: 92%

“…Although plasmin could efficiently process the gp120-gp41 cleavage site, as has been previously reported (61), it also cleaved gp120 at a second site, most probably within the V3 loop. This renders its use impractical.…”

Section: Discussionsupporting

confidence: 64%

“…In principle, one way to achieve Env cleavage is to treat purified Env proteins in vitro with proteases capable of recognizing the gp120-gp41 cleavage site. The highly active subtilisin family protease plasmin was previously reported to cleave recombinant gp160 into gp120-gp41, whereas other trypsin-like proteases lacked this ability (61). Plasmin is also capable of processing influenza virus HA 0 at the cell surface (32).…”

Section: Resultsmentioning

confidence: 99%

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Enhancing the Proteolytic Maturation of Human Immunodeficiency Virus Type 1 Envelope Glycoproteins

Binley

Sanders

Master

et al. 2002

J Virol

143

165

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In virus-infected cells, the envelope glycoprotein (Env) precursor, gp160, of human immunodeficiency virus type 1 is cleaved by cellular proteases into a fusion-competent gp120-gp41 heterodimer in which the two subunits are noncovalently associated. However, cleavage can be inefficient when recombinant Env is expressed at high levels, either as a full-length gp160 or as a soluble gp140 truncated immediately N-terminal to the transmembrane domain. We have explored several methods for obtaining fully cleaved Env for use as a vaccine antigen. We tested whether purified Env could be enzymatically digested with purified protease in vitro. Plasmin efficiently cleaved the Env precursor but also cut at a second site in gp120, most probably the V3 loop. In contrast, a soluble form of furin was specific for the gp120-gp41 cleavage site but cleaved inefficiently. Coexpression of Env with the full-length or soluble form of furin enhanced Env cleavage but also reduced Env expression. When the Env cleavage site (REKR) was mutated in order to see if its use by cellular proteases could be enhanced, several mutants were found to be processed more efficiently than the wild-type protein. The optimal cleavage site sequences were RRRRRR, RRRRKR, and RRRKKR. These mutations did not significantly alter the capacity of the Env protein to mediate fusion, so they have not radically perturbed Env structure. Furthermore, unlike that of wild-type Env, expression of the cleavage site mutants was not significantly reduced by furin coexpression. Coexpression of Env cleavage site mutants and furin is therefore a useful method for obtaining high-level expression of processed Env.The Env glycoprotein complex mediates receptor binding and membrane fusion during human immunodeficiency virus type 1 (HIV-1) infection of susceptible cells (66). It is synthesized as a polypeptide precursor (gp160) that oligomerizes to form a heavily glycosylated trimer (20,24). At a late stage of synthesis, most probably in the trans-Golgi network (TGN), gp160 is cleaved by furin (17,18,(55)(56)(57)(58) or other, related subtilisin-like proteases (17,18,28,38,58,90) into the surface (SU; gp120) and transmembrane (TM; gp41) subunits (34,43,(55)(56)(57)(58)82). Cleavage occurs at a motif at the gp120-gp41 juncture that contains a basic amino acid tetrad, R-X-(R/K)-R (where X is any amino acid). The gp120 and gp41 proteins then remain noncovalently associated, forming the functional, native gp120 3 -gp41 3 complex (20,24,66).During fusion, the gp120 protein interacts with the virus receptor and coreceptor on target cells. This triggers conformational changes that lead to the insertion of a hydrophobic fusion peptide, located at the N terminus of gp41, into the target cell membrane (66). Cleavage of gp160 is essential for fusion, since uncleaved gp160 is fusion incompetent (9,33,39,48). Generally, only cleaved Env is incorporated into virions (22), although uncleaved Env can be virion associated (39,48). By analogy with other enveloped viruses such as influenza A virus (5,...

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Section: Resultssupporting

confidence: 92%

Section: Discussionsupporting

confidence: 64%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Enhancing the Proteolytic Maturation of Human Immunodeficiency Virus Type 1 Envelope Glycoproteins

Binley

Sanders

Master

et al. 2002

J Virol

143

165

View full text Add to dashboard Cite

show abstract

“…Several invasive gram-positive and gram-negative bacteria have shown the ability to interact with the host PLG system (6,13,19,37,73), a phenomenon that has been extended to viruses (22,38,60) and parasites (68) (23,41,46,47,69,74). In the case of spirochetes, the PAS was studied with several species of Borrelia and with Treponema denticola and was suggested to have an important role during infectiveness (14,16,21,35,58).…”

Section: Discussionmentioning

confidence: 99%

“…It has been shown that several pathogens, including the spirochete Borrelia burgdorferi, bind PLG on the surface and convert it to plasmin by host activators (6,13,16,19,22,31,34,37,38,60,68,73); this binding promotes degradation of ECM components and is essential for dissemination of the bacteria through the host tissues, suggesting its role during infection and pathogenesis (12,14,15,28,37,58).…”

mentioning

confidence: 99%

Plasminogen Acquisition and Activation at the Surface of Leptospira Species Lead to Fibronectin Degradation

et al. 2009

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Pathogenic Leptospira species are the etiological agents of leptospirosis, a widespread disease of human and veterinary concern. In this study, we report that Leptospira species are capable of binding plasminogen (PLG) in vitro. The binding to the leptospiral surface was demonstrated by indirect immunofluorescence confocal microscopy with living bacteria. The PLG binding to the bacteria seems to occur via lysine residues because the ligation is inhibited by addition of the lysine analog 6-aminocaproic acid. Exogenously provided urokinase-type PLG activator (uPA) converts surface-bound PLG into enzymatically active plasmin, as evaluated by the reaction with the chromogenic plasmin substrate D-Val-Leu-Lys 4-nitroanilide dihydrochloridein. The PLG activation system on the surface of Leptospira is PLG dose dependent and does not cause injury to the organism, as cellular growth in culture was not impaired. The generation of active plasmin within Leptospira was observed with several nonvirulent high-passage strains and with the nonpathogenic saprophytic organism Leptospira biflexa. Statistically significant higher activation of plasmin was detected with a low-passage infectious strain of Leptospira. Plasmin-coated virulent Leptospira interrogans bacteria were capable of degrading purified extracellular matrix fibronectin. The breakdown of fibronectin was not observed with untreated bacteria. Our data provide for the first time in vitro evidence for the generation of active plasmin on the surface of Leptospira, a step that may contribute to leptospiral invasiveness.

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Furin‐mediated protein processing in infectious diseases and cancer

2019

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Proteolytic cleavage regulates numerous processes in health and disease. One key player is the ubiquitously expressed serine protease furin, which cleaves a plethora of proteins at polybasic recognition motifs. Mammalian substrates of furin include cytokines, hormones, growth factors and receptors. Thus, it is not surprising that aberrant furin activity is associated with a variety of disorders including cancer. Furthermore, the enzymatic activity of furin is exploited by numerous viral and bacterial pathogens, thereby enhancing their virulence and spread. In this review, we describe the physiological and pathophysiological substrates of furin and discuss how dysregulation of a simple proteolytic cleavage event may promote infectious diseases and cancer. One major focus is the role of furin in viral glycoprotein maturation and pathogenicity. We also outline cellular mechanisms regulating the expression and activation of furin and summarise current approaches that target this protease for therapeutic intervention.

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The extracellular processing of HIV‐1 envelope glycoprotein gp160 by human plasmin

Cited by 17 publications

References 24 publications

Enhancing the Proteolytic Maturation of Human Immunodeficiency Virus Type 1 Envelope Glycoproteins

Enhancing the Proteolytic Maturation of Human Immunodeficiency Virus Type 1 Envelope Glycoproteins

Plasminogen Acquisition and Activation at the Surface of Leptospira Species Lead to Fibronectin Degradation

Furin‐mediated protein processing in infectious diseases and cancer

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