Exploring the Molecular Properties of Collagen Type IV with Atomic Force Microscopy

Collagen type IV is a network-forming collagen that provides support and anchorage to cells. Its basic structural unit is a 410 nm long and 1.5 nm in diameter triple helix, with natural discontinuities in the triple-helical defining Gly-X-Y sequence. The C-terminal globular domain (NC1) in a collagen IV molecule plays an important role in forming networks, and has recently been reported to be structurally triggered by chloride ions to form hexamers outside the cell. How this hexamer assembles in vitro remains unknown. Here, I aim to use atomic force microscopy (AFM) to investigate the molecular basis of collagen type IV network assembly by studying the effects of different solvent conditions on the stability of the NC1 domain. Studying the dissociation of this hexametric domain can shed light onto how it assembles in solution and under what ionic conditions. The flexibility of the collagen type IV molecule is also investigated by performing statistical analysis of AFM-imaged chains and estimating persistence length, a mechanical property that quantifies the flexibility of a polymer. Here, I investigate the effects of triple helix interruptions on the flexibility of the molecule, by comparing collagen type IV to other fibrillar collagens that are continuously triple-helical. In addition, I determine a position-dependent flexibility profile of the molecule showcasing the effects of over-lapping interruptions, from a α1(IV)]2–α2(IV) mouse collagen type IV, on the persistence length

Reliability, scalability and robustness issues in iri

Over the past two years, we have used the IRI (Interactive Remote Instruction) system to teach several live interactive classes with students in different cities. While this system is a prototype- we are using it to better understand both system performance requirements and what tools can be effective for remote instruction and how to use them-we have used it repeatedly to teach regularly scheduled forcredit university classes. This repeated use has resulted in significant improvements in IRI’s functionality, but its evaluative use in real classrooms situations has required that we address significant scalability, reliability, and robustness issues. We discuss features of IRI’s software architecture and basic functionality motivated by these scalability and reliability issues. 1

Reliability, Scalability and Robustness Issues in IRI

Over the past two years, we have used the IRI (Interactive Remote Instruction) system to teach several live interactive classes with students in different cities. While this system is a prototype - we are using it to better understand both system performance requirements what tools can be effective for remote instruction, and how to use them - we have used it repeatedly to teach regularly scheduled for-credit university classes. This repeated use has resulted in significant improvements in IRI's functionality, but its evaluative use in real classrooms situations has required that we address significant scalability, reliability, and robustness issues. We discuss features of IRI's software architecture and basic functionality motivated by these scalability and reliability issues. 1 Introduction IRI (Interactive Remote Instruction) is a system under development at Old Dominion University which we are using to support distance learning - education without a central classroom. In this pape..

Sequence-dependent mechanics of collagen reflect its structural and functional organization

Extracellular matrix mechanics influence diverse cellular functions, yet surprisingly little is known about the mechanical properties of their constituent collagen proteins. In particular, network-forming collagen IV, an integral component of basement membranes, has been far less studied than fibril-forming collagens. A key feature of collagen IV is the presence of interruptions in the triple-helix-defining (Gly-X-Y) sequence along its collagenous domain. Here, we used atomic force microscopy to determine the impact of sequence heterogeneity on the local flexibility of collagen IV and of the fibril-forming collagen III. Our extracted flexibility profile of collagen IV reveals that it possesses highly heterogeneous mechanics, ranging from semiflexible regions as found for fibril-forming collagens to a lengthy region of high flexibility toward its N-terminus. A simple model in which flexibility is dictated only by the presence of interruptions fit the extracted profile reasonably well, providing insight into the alignment of chains and demonstrating that interruptions, particularly when coinciding in multiple chains, significantly enhance local flexibility. To a lesser extent, sequence variations within the triple helix lead to variable flexibility, as seen along the continuously triple-helical collagen III. We found this fibril-forming collagen to possess a high-flexibility region around its matrix-metalloprotease binding site, suggesting a unique mechanical fingerprint of this region that is key for matrix remodeling. Surprisingly, proline content did not correlate with local flexibility in either collagen type. We also found that physiologically relevant changes in pH and chloride concentration did not alter the flexibility of collagen IV, indicating such environmental changes are unlikely to control its compaction during secretion. Although extracellular chloride ions play a role in triggering collagen IV network formation, they do not appear to modulate the structure of its collagenous domain