Orthogonal Enzyme-Substrate Design Strategy for Discovery of Human Protein Palmitoyltransferase Substrates.

Protein palmitoylation, with more than 5000 substrates, is the most prevalent form of protein lipidation. Palmitoylated proteins participate in almost all areas of cellular physiology and have been linked to several human diseases. Twenty-three zDHHC enzymes catalyze protein palmitoylation with extensive overlap among the substrates of each zDHHC member. Currently, there is no global strategy to delineate the physiological substrates of individual zDHHC enzymes without perturbing the natural cellular pool. Here, we outline a general approach to accomplish this on the basis of synthetic orthogonal substrates that are only compatible with engineered zDHHC enzymes. We demonstrate the utility of this strategy by validating known substrates and use it to identify novel substrates of two human zDHHC enzymes. Finally, we employ this method to discover and explore conserved palmitoylation in a family of host restriction factors against pathogenic viruses, including SARS-CoV-2.

Synthesis, Enzymatic Peptide Incorporation, and Applications of Diazirine-Containing Isoprenoid Diphosphate Analogues.

Prenylated proteins contain C15 or C20 isoprenoids linked to cysteine residues positioned near their C-termini. Here we describe the preparation of isoprenoid diphosphate analogues incorporating diazirine groups that can be used to probe interactions between prenylated proteins and other proteins that interact with them. Studies using synthetic peptides and whole proteins demonstrate that these diazirine analogues are efficient substrates for prenyltransferases. Photo-cross-linking experiments using peptides incorporating the diazirine-functionalized isoprenoids selectively cross-link to several different proteins. These new isoprenoid analogues should be broadly useful in the studies of protein prenylation.

Catalyst Control in Switching the Site Selectivity of C-H Olefinations of 1,2-Dihydroquinolines: An Approach to Positional-Selective Functionalization of Quinolines.

A unique approach to achieve site-selective C-H olefinations exclusively at the C-3- or C-8-positions in the quinoline framework has been developed by catalyst control. Distal C(3)-H functionalization is achieved by using palladium catalysis, whereas proximal C(8)-H functionalization is obtained by employing ruthenium catalysis. Switching the site selectivity within a single substrate directly indicates two diverse pathways, which are operating under the palladium- and ruthenium-catalyzed reaction conditions.

Palladium-Catalyzed, ortho-Selective C-H Halogenation of Benzyl Nitriles, Aryl Weinreb Amides, and Anilides.

A palladium-catalyzed, ortho-selective C-H halogenation methodology is reported herein. The highlight of the work is the highly selective C(sp2)-H functionalization of benzyl nitriles in the presence of activated C(sp3)-H bond, which results in good yields of the halogenated products with excellent regioselectivity. Along with benzyl nitriles, aryl Weinreb amides and anilides have been evaluated for the transformation using aprotic conditions. Mechanistic studies yield interesting aspects with respect to the pathway of the reaction and the directing group abilities.

Product Control using Substrate Design: Ruthenium-Catalysed Oxidative C-H Olefinations of Cyclic Weinreb Amides.

A new class of Weinreb amides has been developed as directing groups for the ruthenium-catalysed regioselective oxidative C-H olefination. The new Weinreb amides successfully inhibit the N-O bond reductive cleavage usually associated with the cationic ruthenium system, thereby keeping intact the synthetic utility of Weinreb amides. Mechanistic studies reveal interesting aspects of the directing group capabilities of Weinreb amides when compared to simple amides of similar structures.

Fujiwara-Moritani Reaction of Weinreb Amides using a Ruthenium-Catalyzed C-H Functionalization Reaction.

The ruthenium-catalyzed Fujiwara-Moritani reaction (oxidative-Heck reaction) of Weinreb amides is reported herein. The reaction affords exclusively ortho-C-H olefination products, has excellent substrate scope and tolerates halogen functionalities, which increase the synthetic utility of the method. A variety of activated olefins as well as styrenes can be employed as coupling partners.

Heteroatom-guided, palladium-catalyzed regioselective C-H functionalization in the synthesis of 3-arylquinolines.

A new approach for the regioselective functionalization of the C-3-position of quinolines is described. The method utilizes heteroatom guided regioselective C-3 palladation followed by arylation via transmetalation with aryl boronic acids to yield 3-aryl-N-acyl-1,2-dihydroquinolines. In a one-pot sequence, N-deacylation followed by aromatization leads to important 3-arylquinolines in good yields.