The p41 splice variant of major histocompatibility complex (MHC) class II-associated invariant chain (Ii) contains a 65 aa segment that binds to the active site of cathepsin L (CatL), a lysosomal cysteine protease involved in MHC class II-restricted antigen presentation. This segment is absent from the predominant form of Ii, p31. Here we document the in vivo significance of the p41-CatL interaction. By biochemical means and electron microscopy, we demonstrate that the levels of active CatL are strongly reduced in bone marrow-derived antigen-presenting cells that lack p41. This defect mainly concerns the mature two-chain forms of CatL, which depend on p41 to be expressed at wild-type levels. Indeed, pulse-chase analysis suggests that these mature forms of CatL are degraded by endocytic proteases when p41 is absent. We conclude that p41 is required for activity of CatL by stabilizing the mature forms of the enzyme. This suggests that p41 is not merely an inhibitor of CatL enzymatic activity, but serves as a chaperone to help maintain a pool of mature enzyme in late-endocytic compartments of antigen-presenting cells.

Lysosomal cysteine proteases, a subgroup of the cathepsin family, are critical for normal cellular functions such as general protein turnover, antigen processing and bone remodeling. In the past decade, the number of identified human cathepsins has more than doubled and their known role in several pathologies has expanded rapidly. Increased understanding of the structure and mechanism of this class of enzymes has brought on a new fervor in the design of small molecule inhibitors with the hope of producing specific, therapeutic drugs for diseases such as arthritis, allergy, multiple sclerosis, atherosclerosis, Alzheimer's disease and cancer.

Course-based Undergraduate Research Experiences (CUREs) can be a very effective means to introduce a large number of students to research. CUREs are often an extension of the instructor's research, which may make them difficult to replicate in other settings because of differences in expertise or facilities. The BASIL (Biochemistry Authentic Scientific Inquiry Lab) CURE has evolved over the past 4 years as faculty members with different backgrounds, facilities, and campus cultures have all contributed to a robust curriculum focusing on enzyme function prediction that is suitable for implementation in a wide variety of academic settings.

Campus shutdowns during the SARS-CoV-2 pandemic posed unique challenges to faculty and students engaged in laboratory courses. Formerly hands-on experiments had to be quickly pivoted to emergency remote learning. While some resources existed prior to this period, many currently available online modules and/or simulations focus on a single technique. The Biochemistry Authentic Scientific Inquiry Lab (BASIL) curriculum has, for several years, provided a robust, linked, holistic inquiry experience that allows students to make connections between multiple techniques, both computational in nature as well as wet-lab-based. As a course-based undergraduate research experience (CURE), this flexible, module-based curriculum allows students to generate original hypotheses based on analysis of proteins of unknown function. We have taught this curriculum as the upper-level laboratory course on our campuses and were obliged to transition to remote instruction at various points in the course sequence. We report on the experiences of faculty and students over the transition period in this course. Additionally, we report as a case study results of one of our campus’ ongoing discipline-based education research (DBER) on the BASIL curriculum prior to and during remote delivery.

Three-dimensional structures of SARS-CoV-2 and other coronaviral proteins archived in the Protein Data Bank were used to analyze viral proteome evolution during the first six months of the COVID-19 pandemic. Analyses of spatial locations, chemical properties, and structural and energetic impacts of the observed amino acid changes in >48,000 viral proteome sequences showed how each one of the 29 viral study proteins have undergone amino acid changes. Structural models computed for every unique sequence variant revealed that most substitutions map to protein surfaces and boundary layers with a minority affecting hydrophobic cores. Conservative changes were observed more frequently in cores versus boundary layers/surfaces. Active sites and protein-protein interfaces showed modest numbers of substitutions. Energetics calculations showed that the impact of substitutions on the thermodynamic stability of the proteome follows a universal bi-Gaussian distribution. Detailed results are presented for six drug discovery targets and four structural proteins comprising the virion, highlighting substitutions with the potential to impact protein structure, enzyme activity, and functional interfaces. Characterizing the evolution of the virus in three dimensions provides testable insights into viral protein function and should aid in structure-based drug discovery efforts as well as the prospective identification of amino acid substitutions with potential for drug resistance.