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High-speed atomic force microscopy reveals strongly polarized movement of clostridial collagenase along collagen fibrils.

ABSTRACT: Bacterial collagenases involved in donor infection are widely applied in many fields due to their high activity and specificity; however, little is known regarding the mechanisms by which bacterial collagenases degrade insoluble collagen in host tissues. Using high-speed atomic force microscopy, we simultaneously visualized the hierarchical structure of collagen fibrils and the movement of a representative bacterial collagenase, Clostridium histolyticum type I collagenase (ColG), to determine the relationship between collagen structure and collagenase movement. Notably, ColG moved ~14.5?nm toward the collagen N terminus in ~3.8?s in a manner dependent on a catalytic zinc ion. While ColG was engaged, collagen molecules were not only degraded but also occasionally rearranged to thicken neighboring collagen fibrils. Importantly, we found a similarity of relationship between the enzyme-substrate interface structure and enzyme migration in collagen-collagenase and DNA-nuclease systems, which share a helical substrate structure, suggesting a common strategy in enzyme evolution.

SUBMITTER: Watanabe-Nakayama T

PROVIDER: S-EPMC4931465 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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High-speed atomic force microscopy reveals strongly polarized movement of clostridial collagenase along collagen fibrils.

Watanabe-Nakayama Takahiro T Itami Masahiro M Kodera Noriyuki N Ando Toshio T Konno Hiroki H

Scientific reports 20160704

Bacterial collagenases involved in donor infection are widely applied in many fields due to their high activity and specificity; however, little is known regarding the mechanisms by which bacterial collagenases degrade insoluble collagen in host tissues. Using high-speed atomic force microscopy, we simultaneously visualized the hierarchical structure of collagen fibrils and the movement of a representative bacterial collagenase, Clostridium histolyticum type I collagenase (ColG), to determine th ...[more]

PMID: 27373458

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Project description:To penetrate host tissues, histotoxic clostridia secrete virulence factors including enzymes to hydrolyze extracellular matrix. Clostridium histolyticum, recently renamed as Hathewaya histolytica, produces two classes of collagenase (ColG and ColH). The high-speed AFM study showed that ColG collagenase moves unidirectionally to plane collagen fibril and rebundles fibril when stalled . The structural explanation of the roles for the tandem collagen-binding segment (CBDs) is illuminated by its calcium-bound crystal structure at 1.9 Å resolution (Rwork = 15.0%; Rfree = 19.6%). Activation may involve calcium-dependent domain rearrangement supported by both small-angle X-ray scattering and size exclusion chromatography. At pCa ≥ 5 (pCa = -log[Ca2+ ]), the tandem CBD adopts an extended conformation that may facilitate secretion from the bacterium. At pCa ≤ 4, the compact structure seen in the crystal structure is adopted. This arrangement positions the two binding surfaces ~ 55 Å apart, and possibly ushers ColG along tropocollagen molecules that allow for unidirectional movement. A sequential binding mode where tighter binding CBD2 binds first could aid in processivity as well. Switch from processive collagenolysis to fibril rearrangement could be concentration dependent. Collagen fibril formation is retarded at 1 : 1 molar ratio of tandem CBD to collagen. Tandem CBD may help isolate a tropocollagen molecule from a fibril at this ratio. At 0.1 : 1 to 0.5 : 1 molar ratios fibril self-assembly was accelerated. Gain of function as a result of gene duplication of CBD for the M9B enzymes is speculated. The binding and activation modes described here will aid in drug delivery design.Accession codesThe full atomic coordinates of the tandem CBD and its corresponding structure factor amplitudes have been deposited in the Protein Data Bank (PDB accession code 5IKU). Small-angle X-ray scattering data and corresponding ab initio models have been submitted to the Small Angle Scattering Biological Data Bank (SASBDB). Accession codes CL2, collagenase module 2, CN2, CP2 are assigned to envelopes for tandem CBD at -log[Ca2+ ] (pCa) 3, 4, 5, and 6, respectively. Accession code DC64 was assigned to the complex of polycystic kidney disease-CBD1-CBD2 with mini-collagen.

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High-speed atomic force microscopy reveals strongly polarized movement of clostridial collagenase along collagen fibrils.

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High-speed atomic force microscopy reveals strongly polarized movement of clostridial collagenase along collagen fibrils.

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