Purification of the 115-kilodalton exoantigen of Cryptococcus neoformans and its recognition by immune sera

Hamilton, Andrew J.; Goodley, J.M.

doi:10.1128/jcm.31.2.335-339.1993

Cited by 25 publications

(19 citation statements)

References 14 publications

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“…Among these antigens, the 36-kDa antigen of C. neoformans may be recognized by MAbs 7H4 and 5G5, as shown by Hamilton et al (19). These workers also reported a 115-kDa exoantigen for C. neoformans that was recognized by immune sera (18). However, we did not identify such an antigen; the antigens extracted by glass beads in the present study were not exoantigens.…”

Section: Discussionsupporting

confidence: 43%

“…However, the presence of antibodies, acting as potent opsonins (32) required for in vitro antifungal activity by natural killer cells (43) and leukocytes (39), suggests that humoral immunity plays an important role in this infection. Other studies have suggested that antibodies also may be important in immunity against C. neoformans (10,18,26).…”

Section: Cryptococcus Neoformans Causes Infection In Individuals Withmentioning

confidence: 99%

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A 77-kilodalton protein of Cryptococcus neoformans, a member of the heat shock protein 70 family, is a major antigen detected in the sera of mice with pulmonary cryptococcosis

et al. 1997

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Heat shock proteins (HSPs) from several pathogenic microbes have been shown to be target molecules of humoral responses as well as cellular immune responses. However, little is known about target molecules in pulmonary cryptococcosis. Western blotting analysis revealed that experimentally induced pulmonary cryptococcosis in (BALB/c ؋ DBA/2)F 1 mice was associated with the appearance of serum antibodies to a 77-kDa protein derived from Cryptococcus neoformans as well as to 18-, 22-, 25-, 36-, and 94-kDa proteins. Since the 77-kDa band also reacted with rabbit polyclonal antibodies against 70-kDa HSP (HSP70) family members, the protein was predicted to be a member of the HSP70 family. We also purified HSP70 directly from a C. neoformans cell extract by Mono Q fast protein liquid chromatography and ATP-agarose affinity column chromatography and showed that it was positive in immunoblot analysis using either serum from C. neoformans-infected mice or rabbit anti-HSP70 antibodies. N-terminal amino acid sequencing of this purified protein confirmed that the 77-kDa protein was a member of the HSP70 protein family. A 66-kDa protein, which coincidentally purified with the HSP70 protein and was identified as a member of the HSP60 family by N-terminal amino acid sequencing, was not reactive with sera from C. neoformans-infected mice. Thus, a protein associated with the HSP70 family and derived from C. neoformans was a major target molecule of the humoral response in murine pulmonary cryptococcosis.

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Section: Discussionsupporting

confidence: 43%

Section: Cryptococcus Neoformans Causes Infection In Individuals Withmentioning

confidence: 99%

A 77-kilodalton protein of Cryptococcus neoformans, a member of the heat shock protein 70 family, is a major antigen detected in the sera of mice with pulmonary cryptococcosis

et al. 1997

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“…SDS-PAGE and isoelectric focusing. The purification of the SODs was monitored via SDS-PAGE (on 10 and 15% polyacrylamide gels and on 4 to 20% Ready gels [Bio-Rad]), as previously described (9,12), with either Coomassie brilliant blue R-250 (10) or silver stain (Bio-Rad). Isoelectric focusing was performed as previously described (12).…”

Section: Methodsmentioning

confidence: 99%

Biochemical comparison of the Cu,Zn superoxide dismutases of Cryptococcus neoformans var. neoformans and Cryptococcus neoformans var. gattii

Hamilton

Holdom

1997

Infect Immun

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Cu,Zn superoxide dismutases (SODs) have been purified to homogeneity from the two varieties of Cryptococcus neoformans, C. neoformans var. neoformans and var. gattii. The N-terminal amino acid sequences of the two enzymes were similar, though not identical, and demonstrated homology with Cu,Zn SODs from other organisms. SOD activity was present in supernatants from stationary-phase cultures of isolates of C. neoformans var. neoformans and was also present from the mid-log phase onwards in cultures of an acapsular mutant of C. neoformans var. neoformans. SOD activity was practically undetectable in culture supernatants from isolates of C. neoformans var. gattii. The C. neoformans var. neoformans SOD had a reduced relative molecular mass of 19 kDa, and in its nonreduced form the enzyme was present as a 125-kDa species. Isoelectric focusing indicated that four species with pIs of 5.9, 6.15, 6.35, and 6.6 were present. The equivalent reduced molecular mass of the C. neoformans var. gattii enzyme was 19 kDa, with a single species present under nonreducing conditions (relative molecular mass of 145 kDa) with a pI of 7.5. The activities of the enzymes from both varieties were inhibited by KCN; however, the copper chelator diethyldithiocarbamate was inhibitory only against the C. neoformans var. gattii enzyme, as was sodium azide. The C. neoformans var. neoformans SOD was not affected by preincubation for 1 h at 70 degrees C, and it also retained most of its activity when incubated at 37 degrees C relative to its activity when incubated at 20 degrees C, in contrast to the C. neoformans var. gattii enzyme. The pronounced differences in the physical and biochemical characteristics of the Cu,Zn SODs from the two Cryptococcus varieties complement recent reports illustrating the biochemical and genetic differences between C. neoformans var. neoformans and C. neoformans var. gattii, and the successful purification of the two enzymes comprises the first step in determining what role, if any, the cryptococcal Cu,Zn SODs might have in protection against externally generated superoxide.

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“…Recently, there has been renewed interest in antibody immunity to glucuronoxylomannan (GXM) for prevention [19] and treatment [20] of human infection. To date, some studies of C. neoformans protein have been described [21][22][23]. However, little is known about the proteins secreted or released by C. neoformans, despite evidence that they elicit important immune responses.…”

Section: Introductionmentioning

confidence: 99%

Heat shock protein 70 (hsp70) as a major target of the antibody response in patients with pulmonary cryptococcosis

Kakeya

Udono

Maesaki

et al. 1999

Clinical and Experimental Immunology

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Cryptococcus neoformans causes infection in individuals with defective T cell function, such as AIDS, as well as without underlying disease. It has been suggested that humoral as well as cellular immunity might play an important role in the immune response to C. neoformans infection. We have recently shown, using immunoblotting, that the 70-kD hsp family of C. neoformans was the major target molecule of the humoral response in murine pulmonary cryptococcosis. In this study we also used immunoblotting to define the antibody responses in the sera of 24 patients with pulmonary cryptococcosis: 21 proven and three suspected diagnoses. Anti-C. neoformans hsp70 antibody was detected in 16 of 24 (66.7%) patients with pulmonary cryptococcosis. Fourteen of 17 (82.3%) patients with high antigen titres (> or = 1:8) and two of seven (28.6%) patients with low titres (< or = 1:4) had detectable levels of anti-hsp70 antibody. Sera from patients positive for anti-hsp70 antibody showed high titres in the Eiken latex agglutination test for the detection of serum cryptococcal antigen. Our results indicate that the 70-kD hsp family from C. neoformans appears to be a major target molecule of the humoral response, not only in murine pulmonary cryptococcosis, but also in human patients with pulmonary cryptococcosis.

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Purification of the 115-kilodalton exoantigen of Cryptococcus neoformans and its recognition by immune sera

Cited by 25 publications

References 14 publications

A 77-kilodalton protein of Cryptococcus neoformans, a member of the heat shock protein 70 family, is a major antigen detected in the sera of mice with pulmonary cryptococcosis

A 77-kilodalton protein of Cryptococcus neoformans, a member of the heat shock protein 70 family, is a major antigen detected in the sera of mice with pulmonary cryptococcosis

Biochemical comparison of the Cu,Zn superoxide dismutases of Cryptococcus neoformans var. neoformans and Cryptococcus neoformans var. gattii

Heat shock protein 70 (hsp70) as a major target of the antibody response in patients with pulmonary cryptococcosis

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