Elsevier

Meat Science

Volume 52, Issue 1, May 1999, Pages 9-18
Meat Science

Hind-limb protein metabolism and calpain system activity influence post-mortem change in meat quality in lamb

https://doi.org/10.1016/S0309-1740(98)00143-0Get rights and content

Abstract

This study is concerned with the rate of protein turnover in the hind limb muscle bed of intact lambs, the activity of calpain proteolytic system in the M. biceps femoris, and subsequent rates of myofibre breakdown and tenderisation in the M. longissimus dorsi. Feed restriction increased protein degradation in hind-limb muscle of lambs (p < 0.1), with a concominant decrease in the extractable activity of calpastatin (p < 0.01), the endogenous inhibitor of calpain. IGF-1 analog treatment decreased both protein degradation and assayed μ-calpain activity (p < 0.05) with no effect on the activity of calpastatin. β-Agonist treatment increased hind-limb protein synthesis (p < 0.01), calpastatin activity (p < 0.1) and decreased (p < 0.01) μ-calpain activity, but did not effect protein degradation. Significant correlations were observed between Myofibril Fragmentation Index (MFI) values during post-mortem storage and initial post-slaughter calpastatin activity at days 3 (r=−0.34, p < 0.1), 5 (r=−0.58, p < 0.01) and 9 (r=−0.58, p < 0.1), and μ-calpain activity at days 5 (r=0.35, p < 0.1) and 9 (r=0.41, p < 0.05). However, stronger correlations were observed between the ratio of μ-calpain to calpastatin, an estimate of potential μ-calpain proteolytic activity, and the rate of myofibril fragmentation (r=0.75, p < 0.001) and tenderisation (r=−0.64, p < 0.01) during aging. These results are consistent with the calpain system being the major proteolytic system involved in myofibril fragmentation and hence aging-related tenderisation of meat.

Introduction

It is well established that the proteolysis of myofibrillar proteins by endogenous proteases during post-mortem aging is primarily responsible for the tenderisation of meat (Penny, 1980). The major proteins associated with myofibrillar contraction, actin, myosin and α-actinin, are not degraded during the aging process. Tenderisation is therefore thought to result from the cleavage of key structural cytoskeletal proteins such as the intermediate filament and costamere proteins (Taylor et al., 1995). Of the known endogenous proteolytic systems, the calcium activated proteolytic (calpain) system (E.C. 3.4.22.17) is thought to be responsible for the specific cleavage of these proteins during aging (Koohmaraie, 1994). It is becoming increasingly clear that calpains constitute a large family of enzymes (Suzuki et al., 1995). At present, skeletal muscle is known to contain the ubiquitous μ- and m-calpains, active at micro- and milli-molar Ca2+ concentrations, respectively, the muscle-specific p94 calpain, and the calpain-specific inhibitor calpastatin.

Apart from their effects on muscle during post-mortem storage, calpains play an important physiological role in intracellular protein degradation. The calpain system was first implicated in initiating metabolic turnover of myofibrillar proteins by Dayton et al., 1975. Since then, considerable evidence has been accumulated to support the involvement of calpains in skeletal muscle growth and development (Goll et al., 1991, Goll et al., 1992). Lysosomal proteolytic mechanisms account for approximately 25–30% of intracellular protein degradation (Reeves et al., 1981, Lowell et al., 1986), which corresponds to the levels of sarcoplasmic protein in muscle cells (Goll et al.). Also, although the proteasome is thought to be responsible for much of the degradation of myofibrillar proteins to their constituent amino acids through the ubiquitin system (Ciechanover, 1995), it is unable to initiate the disassembly of myofibrillar proteins from the myofibre (Koohmaraie, 1992). Accordingly, it is not unreasonable to believe that if calpains are involved in initiation of protein degradation in muscle in vivo, they may be altered in concert by factors known also to alter protein degradation.

The purpose of this study was to determine associations between variation in protein degradation in muscle of live animals, differences in calpain system activity and the rate of meat tenderisation during aging. Experimental treatments used to alter muscle protein degradation were level of nutrition and two endocrine regulators (clenbuterol, a β-agonist, and Long R3IGF-1, an analog of insulin-like growth factor 1) previously shown to alter muscle protein degradation (Bohorov et al., 1987, Oddy and Owens, 1996).

Section snippets

Animals and experimental design

Twenty six Dorset–Merino–Border Leister-cross castrate lambs with an average weight of 27.0 ± 3.3 kg (s.e.) were used in this study. The experiment was based on a two by three factorial design. Lambs were divided into two groups and fed either at a high or low plane of nutrition. At each plane, animals were subdivided randomly into three groups of at least four lambs for endocrine (control, β-agonist, or IGF-1 analog) treatment.

All lambs were fed a high quality pelleted diet (Thompson et al., 1985

Protein kinetics in hind-limb muscle

No interactions between level of FI and endocrine treatment were observed for hind-limb protein turnover, oxygen consumption and blood flow determinations. Level of FI had no effect on hind-limb arterial–venous oxygen difference, oxygen uptake or blood flow through the infused hind-limb (Table 1). However, nutritional restriction reduced the difference in phenylalanine concentration between the arterial and venous bloods (p < 0.1), and had significant effects on hind-limb protein turnover.

Discussion

It is clear that nutritional and endocrine influences play an important role in regulating protein turnover (Reeds, 1989). This work describes the influences of short-term nutritional restriction, IGF-1 analog and clenbuterol treatments on the rate of protein turnover in the hind limb, levels of calpain system activity, and tenderisation (rate of proteolysis during aging). These observations are more striking given that rate of protein turnover was measured in the entire hind-limb muscle bed,

Acknowledgements

The authors are grateful for the expert technical assistance provided by C. Quinn, R. Woodgate and K. Zirkler. The Long-R3-IGF-1 was kindly donated by Dr. P.C. Owens, Department of Obstetrics and Gynecology, School of Medicine, University of Adelaide, Australia. The authors wish to acknowledge the financial support from the Cattle and Beef Industry Co-operative Research Centre (Meat Quality), and from NSW Agriculture. M.B. McDonagh was in receipt of a Meat Quality CRC postgraduate scholarship

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