Study of the affinity between the protein kinase PKA and homoarginine-containing peptides derived from kemptide: Free energy perturbation (FEP) calculations.

Protein kinases (PKs) discriminate between closely related sequences that contain serine, threonine, and/or tyrosine residues. Such specificity is defined by the amino acid sequence surrounding the phosphorylatable residue, so that it is possible to identify an optimal recognition motif (ORM) for each PK. The ORM for the protein kinase A (PKA), a well-known member of the PK family, is the sequence RRX(S/T)X, where arginines at the -3 and -2 positions play a key role with respect to the primed phosphorylation site. In this work, differential affinities of PKA for the peptide substrate Kemptide (LRRASLG) and mutants that substitute the arginine residues by the unnatural peptide homoarginine were evaluated through molecular dynamics (MD) and free energy perturbation (FEP) calculations. The FEP study for the homoarginine mutants required previous elaboration of a CHARMM "arginine to homoarginine" (R2B) hybrid topology file which is available in this manuscript as Supporting Information. Mutants substituting the arginine residues by alanine, lysine, and histidine were also considered in the comparison by using the same protocol. FEP calculations allowed estimating the free energy changes from the free PKA to PKA-substrate complex (ΔΔGE→ES ) when Kemptide structure was mutated. Both ΔΔGS→ES values for homoarginine mutants were predicted with a difference below 1 kcal/mol. In addition, FEP correctly predicted that all the studied mutations decrease the catalytic efficiency of Kemptide for PKA. © 2018 Wiley Periodicals, Inc.

Arginine Metabolism Revisited.

Mammalian arginine metabolism is complex due to the expression of multiple enzymes that utilize arginine as substrate and to interactions or competition between specific enzymes involved in arginine metabolism. Moreover, cells may contain multiple intracellular arginine pools that are not equally accessible to all arginine metabolic enzymes, thus presenting additional challenges to more fully understanding arginine metabolism. At the whole-body level, arginine metabolism ultimately results in the production of a biochemically diverse range of products, including nitric oxide, urea, creatine, polyamines, proline, glutamate, agmatine, and homoarginine. Included in this group of compounds are the methylated arginines (e.g., asymmetric dimethylarginine), which are released upon degradation of proteins containing methylated arginine residues. Changes in arginine concentration also can regulate cellular metabolism and function via a variety of arginine sensors. Although much is known about arginine metabolism, elucidation of the physiologic or pathophysiologic roles for all of the pathways and their metabolites remains an active area of investigation, as exemplified by current findings highlighted in this review.

Nitric oxide synthesis capacity, ambulatory blood pressure and end organ damage in a black and white population: the SABPA study.

Nitric oxide (NO) synthesis capacity is determined by the availability of substrate(s) such as L-arginine and the influence of nitric oxide synthase (NOS) inhibitors, asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA). These factors may be important in black South Africans with a very high prevalence of hypertension. We compared ambulatory blood pressure (BP), markers of end organ damage and NO synthesis capacity markers [L-arginine, L-homoarginine, L-citrulline, L-arginine:ADMA, ADMA, SDMA and dimethylarginine (DMA)], between black and white teachers (n = 390). Associations of nighttime BP and markers of end organ damage with NO synthesis capacity markers were also investigated. Although black men and women had higher BP and albumin-to-creatinine ratio (ACR) (all p < 0.001), they also had higher L-arginine, L-homoarginine, L-arginine:ADMA and lower SDMA and DMA levels (all p < 0.05). Only in white men ADMA concentrations associated positively with nighttime systolic blood pressure (R (2) = 0.20, ß = 0.26, p = 0.009), nighttime diastolic blood pressure (R (2) = 0.23, ß = 0.27, p = 0.007), carotid intima media thickness (cIMT) (R (2) = 0.36, ß = 0.22, p = 0.008) and ACR (R (2) = 0.14, ß = 0.32, p = 0.001). Our findings suggest that despite an adverse cardiovascular profile in blacks, their NO synthesis capacity profile seems favourable, and that other factors, such as NO inactivation, may prove to be more important.