SummaryProtein design provides an ultimate test of our knowledge about proteins and allows the creation of novel enzymes for biotechnological applications. While progress has been made in designing proteins that mimic native proteins structurally1–3, it is more difficult to design functional proteins4–8. In comparison to recent successes in designing non-metalloproteins4,6,7,9,10, it is even more challenging to rationally design metalloproteins that reproduce both the structure and function of native metalloenzymes5,8,11–20, since protein metal binding sites are much more varied than non-metal containing sites, in terms of different metal ion oxidation states, preferred geometry and metal ion ligand donor sets. Because of their variability, it has been difficult to predict metal binding site properties in silico, as many of the parameters for metal binding sites, such as force fields are ill-defined. Therefore, the successful design of a structural and functional metalloprotein will greatly advance the field of protein design and our understanding of enzymes. Here, we report a successful, rational design of a structural and functional model of a metalloprotein, nitric oxide reductase (NOR), by introducing three histidines and one glutamate, predicted as ligands in the active site of NOR, into the distal pocket of myoglobin. A crystal structure of the designed protein confirms that the minimized computer model contains a heme/non-heme FeB center that is remarkably similar to that in the crystal structure. This designed protein also exhibits NOR activity. This is the first designed protein that models both the structure and function of NOR, offering insight that the active site glutamate is required for both iron binding and activity. These results show that structural and functional metalloproteins can be rationally designed in silico.

A conserved 2-His-1-Glu metal center, as found in natural non-heme iron-containing enzymes, was engineered into sperm whale myoglobin by replacing Leu29 and Phe43 with Glu and His, respectively (swMb L29E, F43H, H64, called Fe B Mb(-His)). A high resolution (1.65 Å) crystal structure of Cu(II)-CN − -Fe B Mb(-His) was determined, demonstrating that the unique 2-His-1-Glu metal center was successfully created within swMb. The Fe B Mb(-His) can bind Cu, Fe or Zn ions, with both Cu(I)-Fe B Mb(-His) and Fe(II)-Fe B Mb(-His) exhibiting nitric oxide reductase (NOR) activities. Cu dependent NOR activity was significantly higher than that of Fe in the same metal yi-lu@illinois.edu . Δ Present address:

BACKGROUND: Social attachment has been identified as a key antecedent motivating users’ social media involvement. However, there is a scarcity of research investigating whether and how three dimensions of social attachment exert impacts on users’ continuous usage intention of social media. OBJECTIVE: Based on structural equation model analysis, the current research clarifies the relationships between social attachment, affective commitment and social media continuous usage intention, which unveils the underlying mechanism through which three dimensions of social attachment influence users’ continuous usage intention of social media. METHODS: A survey was conducted with 536 informative responses obtained from TikTok public users for hypothesis testing analysis. RESULTS: Results indicate that three dimensions of social attachment (social connections, social dependence and social identity) are all positively related to users’ continuous usage intention of social media. Affective commitment partially mediates the relationship between social attachment and users’ continuous usage intention of social media. CONCLUSIONS: The current research makes an in-depth study about the underlying mechanism whereby social attachment exerts impacts on social media continuous usage intentionand provides several managerial and theoretical implications. Future research directions are discussed as well.

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