Characterization of non-cytosolic hexokinase activity in white skeletal muscle from goldfish (Carassius auratus L.) and the effect of cold acclimation

Abstract

HK (hexokinase) is an enzyme involved in the first step in the glucose metabolism pathway, converting glucose into G6P (glucose 6-phosphate). Owing to the importance of skeletal muscle for fish swimming and acclimation processes, we used goldfish (Carassius auratus L.) white muscle in order to investigate subcellular distribution and kinetics of HK. In this study, we report that HK activity is predominantly localized in the mitochondrial fraction [NC-HK (non-cytosolic HK)] in goldfish white muscle. Studies of the kinetic parameters revealed that the Km (Michaelis-Menten constant) for glucose was 0.41±0.03 mM and that for mannose was 3-fold lower, whereas the affinity for fructose was too low to be measured. The Km for ATP was 0.88±0.05 mM, whereas no activity was observed when either GTP or ITP was used as a phosphate donor. A moderate inhibition (20-40%) was found for ADP and AMP. Similar to mammalian HK, G6P and glucose analogues were able to promote an inhibition of between 85 and 100% of activity. Here, we found that acclimation of goldfish at 5°C promoted a 2.5-fold increase in NC-HK compared with its counterpart acclimated at 25°C. However, cytosolic HK activity was not altered after thermal acclimation. In summary, our results suggest that the goldfish has a constitutive NC-HK that shows some similarities to mammalian HK-II and, curiously, may play a role in the broad metabolic changes required during the cold acclimation process.

Effects of Melissa officinalis L. Essential Oil in Comparison with Anaesthetics on Gill Tissue Damage, Liver Metabolism and Immune Parameters in Sea Bass (Lateolabrax maculatus) during Simulated Live Transport

“…Figure 3 C shows that the HK activity of the fish increased gradually with increasing live transport time. This is similar to previous research results; the present study demonstrated that cold conditions induce an increasing trend in the HK activity of fish [ 54 , 55 , 56 ]. PK activity increased at the highest rate in the CK group, indicating that the stress response of the sea bass in this group was the strongest and that the rate of glycolysis was accelerated to resist the stress response.…”

Transcriptome sequencing reveals the differentially expressed lncRNAs and mRNAs in response to cold acclimation and cold stress in Pomacea canaliculata

“…The hexokinase type 2 gene is involved in the glycolytic pathway and may play an important role in increasing glucose content [46]. Its expression was up-regulated in the cold acclimation group after 24 h at 0 °C, consistent with previous studies in goldfish (Carassius auratus L) [47]. Additionally, the decreased expression of UTP-glucose-1-phosphate uridylyltransferase and glycogenin-1 involved in glycogen synthesis may inhibit metabolic pathways related to synthesis of glucose and glycogen.…”

Cold acclimation increases mitochondrial oxidative capacity without inducing mitochondrial uncoupling in goldfish white skeletal muscle

“…Using a protocol for cold acclimation described previously by our group (dos Santos et al, 2010), we measured the different mitochondrial respiratory states in white muscle mitochondria isolated from warm-acclimated and cold-acclimated goldfish (Fig. 1).…”

“…Fish (20–30 g) were maintained at 25°C on a 12 hour:12 hour light/dark cycle and were generally used within 1–1.5 months of arrival. The acclimation was performed using a refrigeration unit (347 CDG-FANEM®, São Paulo, Brazil) as described previously (dos Santos et al, 2010). Fish were kept at least for three days in the lab before starting the experiment in order to minimize the potential stress caused by transportation.…”

“…Because goldfish are particularly cold tolerant, we decided to use an acclimation protocol that has been recently described by our group (dos Santos et al, 2010) to understand what types of changes occur in the mitochondrial respiratory states and mechanisms that could be affecting ATP synthesis. We compared white muscle mitochondria isolated from goldfish acclimated to 25°C or 5°C for one month.…”