A multi-pronged search for a common structural motif in the secretion signal of Salmonella enterica serovar Typhimurium type III effector proteins

Journal Article · Mon Nov 08 04:00:00 EST 2010 · Molecular Biosystems, 6(12):2448-2458

DOI:https://doi.org/10.1039/c0mb00097c· OSTI ID:994022

Buchko, Garry W; Niemann, George; Baker, Erin Shammel; Belov, Mikhail E; Smith, Richard D; Heffron, Fred; Adkins, Joshua N; McDermott, Jason E

Many pathogenic Gram-negative bacteria use a type III secretion system (T3SS) to deliver effector proteins into the host cell where they reprogram host defenses and facilitate pathogenesis. While it has been determined that the first 20 - 30 N-terminal residues usually contain the ‘secretion signal’ that targets effector proteins for translocation, the molecular basis for recognition of this signal is not understood. Recent machine-learning approaches, such as SVM-based Identification and Evaluation of Virulence Effectors (SIEVE), have improved the ability to identify effector proteins from genomics sequence information. While these methods all suggest that the T3SS secretion signal has a characteristic amino acid composition bias, it is still unclear if the amino acid pattern is important and if there are any unifying structural properties that direct recognition. To address these issues a peptide corresponding to the secretion signal for Salmonella enterica serovar Typhimurium effector SseJ was synthesized (residues 1-30, SseJ) along with scrambled peptides of the same amino acid composition that produced high (SseJ-H) and low (SseJ-L) SIEVE scores. The secretion properties of these three peptides were tested using a secretion signal-CyaA fusion assay and their structures systematically probed using circular dichroism, nuclear magnetic resonance, and ion mobility spectrometry-mass spectrometry. The signal-CyaA fusion assay showed that the native and SseJ-H fusion constructs were secreted into J774 macrophage at similar levels via the SPI-2 secretion pathway while secretion of the SseJ-L fusion construct was substantially retarded, suggesting that the SseJ secretion signal was sequence order dependent. The structural studies showed that the SseJ, SseJ-H, and SseJ-L peptides were intrinsically disordered in aqueous solution with only a small predisposition to adopt nascent helical structure in the presence of the powerful structure stabilizing agent, 1,1,1,3,3,3-hexafluoroisopropanol (HFIP). Intrinsic disorder may be a universal feature of effector secretion signals as analogous conclusions were reached following a similar structural characterization of peptides corresponding to the N-terminal regions of the S. Typhimurium effectors SptP, SopD-2, GtgE, and the Yersinia pestis effector YopH.

• Research Organization: Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)
• Sponsoring Organization: USDOE
• DOE Contract Number: AC05-76RL01830
• OSTI ID: 994022
• Report Number(s): PNNL-SA-73691; 40072; 42294; 35407; 33210; 32707; 400412000
• Journal Information: Molecular Biosystems, 6(12):2448-2458, Journal Name: Molecular Biosystems, 6(12):2448-2458 Journal Issue: 12 Vol. 6
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
99 GENERAL AND MISCELLANEOUS
AMINO ACIDS
AQUEOUS SOLUTIONS
BACTERIA
DICHROISM
EVALUATION
Environmental Molecular Sciences Laboratory
ION MOBILITY
MACROPHAGES
NMR spectroscopy
NUCLEAR MAGNETIC RESONANCE
PATHOGENESIS
PEPTIDES
PROTEINS
RESIDUES
SALMONELLA
SECRETION
SPECTROSCOPY
TARGETS
TRANSLOCATION
VIRULENCE
circular dichroism
infectious diseases
ion mobility spectrometry-mass spectrometry
structural biology