Changes in tissue abundance and activity of enzymes related to branched-chain amino acid catabolism in dairy cows during early lactation

Abstract

Branched-chain α-keto acid dehydrogenase (BCKDH) complex catalyzes the irreversible oxidative decarboxylation of branched-chain α-keto acids. This reaction is considered as the rate-limiting step in the overall branched-chain amino acid (BCAA) catabolic pathway in mammals. For characterizing the potential enzymatic involvement of liver, skeletal muscle, adipose tissue (AT), and mammary gland (MG) in BCAA metabolism during early lactation, tissue and blood samples were examined on d 1, 42, and 105 after parturition from 25 primiparous Holstein cows. Serum BCAA profiles were analyzed and the mRNA and protein abundance as well as the activity in the different tissues were assessed for the BCAA catabolic enzymes, partly for the branched-chain aminotransferase and completely for BCKDH. Total BCAA concentration in serum was lowest on d 1 after parturition and increased thereafter to a steady level for the duration of the experiment. Pronounced differences between the tissues were observed at all molecular levels. The mRNA abundance of the mitochondrial isoform of branched-chain aminotransferase (BCATm) was greatest in AT as compared with the other tissues studied, indicating that AT might be an important contributor in the initiation of BCAA catabolism in dairy cows. From the different subunits of the BCKDH E1 component, only the mRNA for the β polypeptide (BCKDHB), not for the α polypeptide (BCKDHA), was elevated in liver. The BCKDHA mRNA abundance was similar across all tissues except muscle, which tended to lower values. Highest BCK-DHA protein abundance was observed in both liver and MG, whereas BCKDHB protein was detectable in these tissues but could not be quantified. Adipose tissue and muscle only displayed abundance of the α subunit, with muscle having the lowest BCKDHA protein of all tissues. We found similarities in protein abundance for both BCKDH E1 subunits in liver and MG; however, the corresponding overall BCKDH enzyme activity was 7-fold greater in liver compared with MG, allowing for hepatic oxidation of BCAA transamination products. Reduced BCKDH activity in MG associated with no measurable activity in AT and muscle may favor sparing of BCAA for the synthesis of the different milk components, including nonessential AA. Deviating from previously published data on BCAA net fluxes and isotopic tracer studies in ruminants, our observed results might in part be due to complex counter-regulatory mechanisms during early lactation.

Branched-chain amino acids: Abundance of their transporters and metabolizing enzymes in adipose tissue, skeletal muscle, and liver of dairy cows at high or normal body condition

“…It has been acknowledged though, that the initial step in BCAA catabolism, the reversible deamination via BCAT mostly occurs in extrahepatic tissues, such as muscle and AT (Bergen et al, 1988;Lapierre et al, 2002;Zhang et al, 2017). Consequently, and in support of previously published data (Suryawan et al, 1998;Webb et al, 2019), we detected the lowest BCAT2 mRNA abundance in liver. This may be indicative of a reduced hepatic BCAA transamination capacity, allowing for a greater availability of BCAA to peripheral tissues regardless of differences in DMI.…”

“…Protein extraction and quantification via Simple Western size-based protein assay (WES, ProteinSimple, San Jose, CA) were described earlier (Webb et al, 2019). In brief, after tissue homogenization, samples were diluted with 0.1 × sample buffer to a protein concentration of 0.325 mg/mL for liver and 0.5 mg/ mL for muscle and scAT, respectively, and then mixed with 5 × master mix containing 40 mM dithiothreitol.…”

“…In contrast to most other EAA, only a small proportion of absorbed BCAA is directly removed by the ruminant liver (Lapierre et al, 2002;Raggio et al, 2004) and at least the initial catabolic step, the reversible deamination via the branched-chain aminotransferase (BCAT; EC 2.6.1.42), yielding the respective branched-chain α-keto acids (BCKA; ketoisoleucine, ketoleucine, ketovaline) and glutamate, may occur extrahepatically [e.g., in muscle (Bequette et al, 2002) and AT (Bergen et al, 1988)]. However, the subsequent oxidative decarboxylation of the BCKA to branched-chain acyl CoA derivatives, catalyzed by the rate-limiting branched-chain α-keto acid dehydrogenase (BCKDH; EC 1.2.4.4) complex, may take place in liver again to maintain physiological BCAA/BCKA levels (Ananieva et al, 2017;Webb et al, 2019). Whole-body BCAA homeostasis is therefore highly dependent on the crosstalk between various tissues.…”

“…Different studies have shown that AT may be an important site for BCAA metabolism (Bergen et al, 1988;Herman et al, 2010;Liang et al, 2019;Webb et al, 2019), in some cases even during periods of negative energy balance. Regardless of this however, a possible link between BCAA degradation and fatty acid metabolism, in particular glyceroneogenesis and fatty acid oxidation, might exist via the tricarboxylic acid (TCA) cycle (Kainulainen et al, 2013).…”

Analysis of Long Non-Coding RNAs and mRNAs Associated with Lactation in the Crop of Pigeons (Columba livia)

“…In the propanoate metabolism pathway, the up-regulated genes promoted the synthesis of lipids in the lactating crop. BCKDH participated in the branched-chain amino acid catabolic pathway by catalyzing the oxidative decarboxylation of the branched-chainα-keto acids [38]. ACAT1 and ACAT2 were acetyl-CoA acetyltransferases, corresponding to two enzymes localized in the mitochondria and cytoplasm respectively.…”