scholarly journals Structural determinants of the interaction between the Haemophilus influenzae Hap autotransporter and fibronectin

Microbiology ◽  
2014 ◽  
Vol 160 (6) ◽  
pp. 1182-1190 ◽  
Author(s):  
Nicole A. Spahich ◽  
Roma Kenjale ◽  
Jessica McCann ◽  
Guoyu Meng ◽  
Tomoo Ohashi ◽  
...  

Haemophilus influenzae is a Gram-negative cocco-bacillus that initiates infection by colonizing the upper respiratory tract. Hap is an H. influenzae serine protease autotransporter protein that mediates adherence, invasion and microcolony formation in assays with human epithelial cells and is presumed to facilitate the process of colonization. Additionally, Hap mediates adherence to fibronectin, laminin and collagen IV, extracellular matrix (ECM) proteins that are present in the respiratory tract and are probably important targets for H. influenzae colonization. The region of Hap responsible for adherence to ECM proteins has been localized to the C-terminal 511 aa of the Hap passenger domain (HapS). In this study, we characterized the structural determinants of the interaction between HapS and fibronectin. Using defined fibronectin fragments, we established that Hap interacts with the fibronectin repeat fragment called FNIII(1–2). Using site-directed mutagenesis, we found a series of motifs in the C-terminal region of HapS that contribute to the interaction with fibronectin. Most of these motifs are located on the F1 and F3 faces of the HapS structure, suggesting that the F1 and F3 faces may be responsible for the HapS–fibronectin interaction.

2002 ◽  
Vol 70 (9) ◽  
pp. 4902-4907 ◽  
Author(s):  
Doran L. Fink ◽  
Bruce A. Green ◽  
Joseph W. St. Geme

ABSTRACT Nontypeable Haemophilus influenzae (NTHI) initiates infection by colonizing the upper respiratory tract mucosa. NTHI disease frequently occurs in the context of respiratory tract inflammation, where organisms encounter damaged epithelium and exposed basement membrane. In this study, we examined interactions between the H. influenzae Hap adhesin and selected extracellular matrix proteins. Hap is an autotransporter protein that undergoes autoproteolytic cleavage, with release of the adhesive passenger domain, Haps, from the bacterial cell surface. We found that Hap promotes bacterial adherence to purified fibronectin, laminin, and collagen IV and that Hap-mediated adherence is enhanced by inhibition of autoproteolysis. Adherence is inhibited by pretreatment of bacteria with a polyclonal antiserum recognizing Haps. Purified Haps binds with high affinity to fibronectin, laminin, and collagen IV but not to collagen II. Binding of Haps to fibronectin involves interaction with the 45-kDa gelatin-binding domain but not the 30-kDa heparin-binding domain of fibronectin. Taken together, these observations suggest that interactions between Hap and extracellular matrix proteins may play an important role in NTHI colonization of the respiratory tract.


2009 ◽  
Vol 77 (11) ◽  
pp. 4704-4713 ◽  
Author(s):  
Roma Kenjale ◽  
Guoyu Meng ◽  
Doran L. Fink ◽  
Twyla Juehne ◽  
Tomoo Ohashi ◽  
...  

ABSTRACT Haemophilus influenzae is a gram-negative bacterium that initiates infection by colonizing the upper respiratory tract. The H. influenzae Hap autotransporter protein mediates adherence, invasion, and microcolony formation in assays with respiratory epithelial cells and presumably facilitates colonization. The serine protease activity of Hap is associated with autoproteolytic cleavage and extracellular release of the HapS passenger domain, leaving the Hapβ C-terminal domain embedded in the outer membrane. Cleavage occurs most efficiently at the LN1036-37 peptide bond and to a lesser extent at three other sites. In this study, we utilized site-directed mutagenesis, homology modeling, and assays with a peptide library to characterize the structural determinants of Hap proteolytic activity and cleavage specificity. In addition, we used homology modeling to predict the S1, S2, and S4 subsite residues of the Hap substrate groove. Our results indicate that the P1 and P2 positions at the Hap cleavage sites are critical for cleavage, with leucine preferred over larger hydrophobic residues or other amino acids in these positions. The substrate groove is formed by L263 and N274 at the S1 subsite, R264 at the S2 subsite, and E265 at the S4 subsite. This information may facilitate design of approaches to block Hap activity and interfere with H. influenzae colonization.


2015 ◽  
Vol 213 (10) ◽  
pp. 1596-1605 ◽  
Author(s):  
Joseph A. Lewnard ◽  
Noga Givon-Lavi ◽  
Amit Huppert ◽  
Melinda M. Pettigrew ◽  
Gili Regev-Yochay ◽  
...  

mSphere ◽  
2019 ◽  
Vol 4 (6) ◽  
Author(s):  
Ilke De Boeck ◽  
Stijn Wittouck ◽  
Katleen Martens ◽  
Jos Claes ◽  
Mark Jorissen ◽  
...  

ABSTRACT It is generally believed that the microbiome plays a role in the pathophysiology of chronic rhinosinusitis (CRS), though its exact contribution to disease development and severity remains unclear. Here, samples were collected from the anterior nares, nasopharynx, and maxillary and ethmoid sinuses of 190 CRS patients and from the anterior nares and nasopharynx of 100 controls. Microbial communities were analyzed by Illumina sequencing of the V4 region of 16S rRNA. The phenotype and patient characteristics were documented, and several serum inflammatory markers were measured. Our data indicate a rather strong continuity for the microbiome in the different upper respiratory tract (URT) niches in CRS patients, with the microbiome in the anterior nares being most similar to the sinus microbiome. Bacterial diversity was reduced in CRS patients without nasal polyps compared to that in the controls but not in CRS patients with nasal polyps. Statistically significant differences in the presence/absence or relative abundance of several taxa were found between the CRS patients and the healthy controls. Of these, Dolosigranulum pigrum was clearly more associated with URT samples from healthy subjects, while the Corynebacterium tuberculostearicum, Haemophilus influenzae/H. aegyptius, and Staphylococcus taxa were found to be potential pathobionts in CRS patients. However, CRS versus health as a predictor explained only 1 to 2% of the variance in the microbiome profiles in an adonis model. A history of functional endoscopic sinus surgery, age, and sex also showed a minor association. This study thus indicates that functional studies on the potential beneficial versus pathogenic activity of the different indicator taxa found here are needed to further understand the pathology of CRS and its different phenotypes. (This study has been registered at ClinicalTrials.gov under identifier NCT02933983.) IMPORTANCE There is a clear need to better understand the pathology and specific microbiome features in chronic rhinosinusitis patients, but little is known about the bacterial topography and continuity between the different niches of the upper respiratory tract. Our work showed that the anterior nares could be an important reservoir for potential sinus pathobionts. This has implications for the diagnosis, prevention, and treatment of CRS. In addition, we found a potential pathogenic role for the Corynebacterium tuberculostearicum, Haemophilus influenzae/H. aegyptius, and Staphylococcus taxa and a potential beneficial role for Dolosigranulum. Finally, a decreased microbiome diversity was observed in patients with chronic rhinosinusitis without nasal polyps compared to that in healthy controls but not in chronic rhinosinusitis patients with nasal polyps. This suggests a potential role for the microbiome in disease development or progression of mainly this phenotype.


2007 ◽  
Vol 35 (4) ◽  
pp. 554-563 ◽  
Author(s):  
XZ Shen ◽  
Q Lu ◽  
L Deng ◽  
S Yu ◽  
H Zhang ◽  
...  

This prospective, three-centre study tested for antimicrobial susceptibility in 898 isolates of Haemophilus influenzae between 2000 and 2002 in Chinese children aged under 5 years with acute upper respiratory tract infection. The average incidence of β-lactamase production was 12.0%. Overall, 88.0% of isolates were susceptible to ampicillin, 100.0% were susceptible to amoxicillin/clavulanic acid, ceftriaxone, cefuroxime and azithromycin, and 99.0% were susceptible to ciprofloxacin. Isolates from Beijing and Shanghai had a lower susceptibility to tetracycline (57.0% and 61.0%, respectively) compared with those from Guangzhou (81.0%), while trimethoprim/sulfamethoxazole susceptibilities in Shanghai (47.0%) and Guangzhou (54.0%) were significantly higher than in Beijing (35.0%). A total of 34.5% of all the isolates were susceptible to all eight of these antimicrobial agents and 12.8% were multi-drug resistant. Ampicillin resistance increased over the duration of the study. These findings show that β-lactamase production and ampicillin resistance among isolates from Chinese children with upper respiratory tract infection are increasing, and highlight the strong correlation between ampicillin resistance and resistance to cefaclor, chloramphenicol and tetracycline in H. influenzae isolates.


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